Human colorectal carcinoma: Patterns of sensitivity to chemotherapeutic agents in the human tumor stem cell assay

The human tumor stem cell assay (HTSCA) was applied to 103 primary and metastatic colorectal carcinomas. Thirty-four carcinomas could not be evaluated for colony formation (clonogenicity) because of microbial contamination. Of the remaining 69 carcinomas, 18 (26%) demonstrated clonal growth in vitro. Colony formation did not correlate with the clini-copathologic stage of the tumor, the histological grade of the tumor, the method used to disperse the solid tumors into single cells (mechanical or enzymatic), or cell viability (exclusion of trypan-blue) prior to plating. The in vitro chemotherapy sensitivity data of the 18 tumors which formed colonies indicated that most tumors were generally resistant to chemotherapeutic drugs and that a few tumors were sensitive to multiple agents. Use of the human tumor stem cell assay may facilitate an individualized approach to clinical chemotherapy of colorectal carcinoma for certain patients.     

Comparison of the human tumor cloning and subrenal capsule assays

Fresh surgical explants of solid tumors obtained from 84 patients were tested against the same chemotherapeutic agents in both the in vitro human tumor cloning (HTC) assay and the in vivo subrenal capsule (SRC) assay. Control growth adequate to meet evaluable assay criteria was obtained in 75 of 84 tumors tested in the SRC assay (89%) and in 33 of 79 tumors tested in the HTC assay (42%). Correlations between the two test systems were dependent upon the activity criteria established for each system. With activity criteria set at current drug screening levels as a decrease of greater than or equal to 50% in tumor colony-forming units for the HTC assay and a change in tumor size less than -1.0 ocular micrometer unit for the SRC assay, 16% of the drugs tested were active in the SRC assay, and 7% were active in the HTC assay. Correlations of tumor response between the two assays were 29% for sensitive (2 of 7) and 83% for resistant (63 of 76). Of the 84 patients providing tumor tissue for assay, 17 had clinically evaluable disease and received chemotherapy providing information for retrospective analysis. A total of 23 SRC assay-clinical correlations and 10 HTC assay-clinical correlations were possible. The SRC assay was predictive of clinical sensitivity in three of three drug tests (100%) and of clinical resistance in 16 of 20 drug tests (80%). No HTC assay-clinical correlations were possible for sensitivity, but the HTC assay was predictive of clinical resistance in 10 of 10 drug tests (100%).     

Prospective correlative chemosensitivity testing in high-dose intraarterial chemotherapy for liver metastases

Clinical response of liver metastases treated by high-dose intraarterial chemotherapy (HDIAC) delivered via the hepatic artery was predicted by a modification of the human tumor colony-forming assay (HTCFA) originally described by Hamburger and Salmon [Science (Wash. DC), 197:461-463, 1977. In a first set of experiments, the immediate clinical response to HDIAC was determined in 12 patients with colorectal liver metastases. Biopsies were taken immediately before and after HDIAC, and cells were plated in the HTCFA. Three patients received intraoperative 4-epidoxorubicin and another 9 received mitomycin C by 15-min intraarterial infusions. Sensitivity in the HTCFA was defined as 50% inhibition of colony formation in tumors exposed to the chemotherapeutic agent, compared to the untreated controls. Clinical response was accurately predicted by the HTCFA in 11 of 12 cases. Eight patients had a regression of disease following HDIAC treatment with mitomycin C, as evidenced by either greater than 50% reduction in carcinoembryonic antigen serum level (7 patients) or regression of tumor by computed tomography scan (1 patient). Three patients had no evidence of clinical response to epidoxorubicin, and their tumors were resistant to epidoxorubicin in the HTCFA. One tumor was sensitive to mitomycin C in the HTCFA, but serum carcinoembryonic antigen in the patient continued to increase following HDIAC. The HTCFA was also performed on untreated biopsies following incubation in vitro with the drug used for HDIAC. Results correlated with clinical response in all 12 cases. In a second set of experiments, the HTCFA was used to predict the long-term clinical response to HDIAC of 30 patients with liver metastases. One patient had breast cancer metastases, one patient had carcinoid liver metastases, 4 had liver metastases of malignant melanoma, and 24 patients had colorectal liver metastases. All 21 of the patients whose tumors were sensitive in vitro had clinical response, while 6 of 9 patients predicted by the HTCFA to be resistant had no clinical response. Our results demonstrate a high correlation between the HTCFA and clinical response.     

In vitro evaluation of anticancer drugs in relation to development of drug resistance in the human tumor clonogenic assay

Summary The efficacy of anticancer drugs against ovarian cancer, breast cancer, and colorectal cancer has been evaluated in vitro by the human tumor clonogenic assay developed by Hamburger and Salmon. The in vitro colony assay method used in this study is a minor modification of their method and was used in 83 patients with ovarian cancer, 47 patients with breast cancer, and 13 patients with colorectal cancer. The total numbers of assays performed in vitro were 258 for ovarian cancer, 87 for breast cancer, and 38 for colorectal cancer. The average chemosensitivity rates to single agents tested were 35% and 32% in the untreated patients with ovarian and breast cancer, respectivety. In contrast to this result, the chemosensitivity rate of the untreated patients with colorectal cancer was only 16%. Consisting the clinical efficacy of anticancer drugs against these tumors, these results suggest that there is a correlation between chemosensitivity in the human tumor clonogenic assay and clinical responsiveness. In this assay the chemosensitivity in specimens from ovarian cancer patients who had had prior chemotherapy was significantly lower than in those from nonpretreated patients (P0.05). This seems to indicate the development of drug resistance after treatment with anticancer drugs. These results suggest that the human tumor clonogenic assay is a useful tool for the evaluation of antitumor effects of drugs in vitro.     

Unexpected in vitro chemosensitivity of malignant gliomas to 4-hydroxyperoxycyclophosphamide (4-HC)

To individually tailor chemotherapy for patients with malignant gliomas according to tumor chemosensitivity, a rapid assay system which can be performed with a high success rate is needed. The fluorescent cytoprint assay (FCA) can assess multiple chemotherapeutic agents using small ( 500 cells) tumor aggregates very quickly ( 1 wk). Tissue samples from 51 patients with malignant gliomas obtained either at time of initial diagnosis (n=34) or at recurrence were assayed using this method. The assay success rate approached 90% in those culture samples which were histologically verified as tumor. A meaningful number of agents could be tested both on samples obtained by stereotactic biopsy (median, 5) and on specimens from more extensive resections (median, 6). One hundred ninety-three FCAs were performed on a samples obtained from 36 patients. In only twenty six assays (14%) was an agent deemed sensitive (> 90% cell kill) to a chemotherapeutic agent. Sixty-two percent of sensitive FCAs were observed in tumors tested against the activated analog of cyclophosphamide, 4-hydroxyperoxycyclophosphamide (4-HC), where a sensitivity rate (# samples sensitive/total tested against agent) of 64% (95% CI, 36.6–77.9%) was noted. This rate was significantly higher than with any other agent tested (p=0.012, two sided McNemar's test) and was not affected by age, histology or disease status. We conclude that: (1) the FCA represents a feasible method for quickly assaying tumors for sensitivity to multiple chemotherapeutic agents; and (ii) malignant gliomas may be particularly sensitive to 4-HC.     

Human tumor clonogenic assay in human breast cancer

The human tumor clonogenic assay (HTCA) developed by Hamburger and Salmon was evaluated in 135 fresh samples of breast cancer. Successful tumor colony growth (greater than or equal to 5 colonies/plate) was obtained in 100 (74%) of the 135 samples, and adequate growth for drug testing (greater than or equal to 30 colonies/plate) in 75 (56%). With regard to the success rates of growing colonies categorized by specimen source, tumor sites, histology, prior chemotherapy and estrogen receptor (ER), specimens from solid tumors and primary tumors showed higher success rates than those from pleural effusions and metastatic tumors. The effects of prior chemotherapy, histology type and ER status on the success rate of colony formation were not significant. The overall median plating efficiency was 0.012%. Higher plating efficiencies were found in pleural effusion, metastatic tumors and samples from patients with prior chemotherapy. These findings appeared to indicate that aggressiveness of disease might be related to plating efficiency. Defining a greater than or equal to 50% inhibition of colony formation (ICF) as in vitro drug sensitivity, the in vitro response rates to anticancer drugs tested were as follows: adriamycin 33%, mitomycin C 39%, 5-fluorouracil 32%, methotrexate 42%, L-PAM 31%, cisplatin 38%, vincristine 32%, vinblastine 54%. The group of patients without prior chemotherapy showed higher sensitivity in vitro compared with the group of patients who had prior chemotherapy (38% vs. 27%). Correlation between in vitro drug sensitivity (greater than or equal to 70% ICF) and clinical response in 12 patients treated with the same drugs were analyzed retrospectively. The predictive accuracy was 0% (0/1) for true positive and 91% (10/11) for resistance. Thus, overall predictive accuracy was 83%. Based on these results, HTCA appeared to be useful chemosensitivity test for evaluation of antitumor drugs for human cancers in vitro and prediction of the chemotherapeutic effect in clinical use.     

Growth of astrocytomas in the human tumor clonogenic assay and sensitivity to mismatched dsRNA and interferons

Nine astrocytoma specimens were received from seven patients and processed for testing in the human tumor clonogenic assay (HTCA). Cells derived from these specimens were challenged with human natural alpha-interferon (alpha-IFN) and beta interferon (beta-IFN), recombinant beta interferon (beta ser-IFN), and mismatched double-stranded (ds) RNA (Ampligen). Six of the astrocytoma specimens formed adequate colonies for drug sensitivity testing (greater than or equal to 30 colonies/plate), and all were high-grade (III-IV) tumors. Sensitivity was defined as a greater than or equal to 50% decrease in tumor colony formation following drug exposure and was observed with alpha-IFN (2/4), beta-IFN (3/4), and mismatched dsRNA (4/5) exposure. No decrease in colony growth was observed after recombinant beta ser-IFN exposure, and in 2 of 3 cases, colony formation was stimulated. The sensitivity of 75 non-CNS solid tumors to mismatched dsRNA was compared to the high-grade astrocytomas in the HTCA. Of the 10 additional histologic tumor types studied, carcinoid and renal cell carcinomas exhibited the greatest sensitivity to mismatched dsRNA: 63% and 52%, respectively. However, in comparison, 80% of the high-grade astrocytomas were sensitive, demonstrating that these gliomas are among the most sensitive of human tumors to mismatched dsRNA in vitro. Clinical trials of interferons and mismatched dsRNA, coupled with in vitro sensitivity studies, should further define their therapeutic potential.     

Application of a human tumor colony-forming assay to new drug screening

The applicability of a human tumor colony-forming assay to drug screening was investigated in terms of feasibility, validity, and potential for discovering new antitumor drugs. Feasibility was addressed in a pilot study during which basic methods, appropriate assay quality controls, and a standardized protocol for screening were developed. Considerable variability was noted in the availability and colony growth of different tumor types. The majority of the evaluable experiments utilized breast, colorectal, kidney, lung, melanoma, or ovarian tumors. For many tumor types, little evidence of growth was observed, or only rare specimens formed colonies. Colony-forming efficiencies ranged from 0.05 to 0.11% for the six most useful tumors listed above. A set of quality control measures was developed to address technical problems inherent in the assay. Testing of standard agents in the pilot study established that most of these agents could be detected as active. However, it also identified three assay limitations: compounds requiring systemic metabolic activation are inactive; medium constituents may block the activity of certain antimetabolites; and compounds without therapeutic efficacy may be positive in the assay. The assay categorized nontoxic clinically ineffective agents as true negatives with 97% accuracy. Of 79 compounds which were negative in the current National Cancer Institute prescreen (leukemia P388), 14 were active in the assay. Several demonstrated outstanding in vitro activity and are structurally unrelated to compounds already in development or in clinical trials. A subset of these active compounds were found to lack activity in a P388 in vitro colony-forming assay. This indication of differential cytotoxicity to human tumor cells makes this subset of compounds particularly interesting as antitumor drug leads. The demonstrated sensitivity to most standard agents, discrimination of nontoxic compounds, reproducibility of survival values within assays and between laboratories, and evidence of ability to identify active compounds which were negative in the in vivo prescreen suggest that the human tumor colony-forming assay may be a valuable tool for antitumor drug screening. However, because of technical limitations inherent in the current assay methodology, this must be confined to selected tumor types and limited to screening on a moderate scale.     

Chemotherapy responsiveness of human tumors as first transplant generation xenografts in the normal mouse: six-day subrenal capsule assay

Feasibility of utilizing human tumors as first transplant generation xenografts in the normal immunocompetent mouse for determining tumor sensitivity to chemotherapeutic agents was demonstrated by applying subrenal capsule (SRC) assay methodology to fresh surgical explants in a six-day time frame. A total of 37 human breast tumors were tested in assays in which 254 xenografts were implanted into control animals. Fifty (20%) of the controls showed some degree of partial regression in the six-day assay period. Using a mean control growth having a positive change in tumor size as the criterion for evaluability, first transplant generation human breast tumors provided an evaluable assay rate of 86%. A tumor response profile was obtained as a result of testing seven clinically active drugs against 32 previously untreated breast cancers. The pattern of responses obtained indicated that no single agent was active against all tumors, nor were tumors which were responsive to one agent necessarily responsive to another, suggesting the feasibility of predicting individual tumor response to specific chemotherapeutic agents. Had these seven drugs been developmental agents of unknown activity which were being tested for the first time against such a panel of human tumors the result would have not only predicted their clinical activity, but the tumor response rates would have also provided an indication of the relative potential of each drug for the specific treatment of breast cancer.     

The clonogenic assay with human tumor xenografts: evaluation, predictive value and application for drug screening

The feasibility, evaluation and predictive value of the colony-forming assay with human tumor xenografts for screening anticancer drugs have been studied. Using human tumors grown in serial passage in nude mice, adequate colony formation was observed in 215 of 251 (86%) different solid human tumors of various histologies. Based on in vitro growth characteristics, a quality-controlled assay protocol was developed. With the proposed criteria for standardized evaluation of individual experiments a substantial increase in assay reliability was achieved. The five clinically established agents, cisplatin, doxorubicin, etoposide, mitomycin-C and vindesine, were studied for anticancer activity in the clonogenic assay. Drugs were applied over a wide dose range by continuous exposure, yielding clear dose-response effects with coefficients of correlation between r = 0.946 and 0.995. Relevant dose levels predicting correctly for the clinical efficacy of the agents were determined by comparison of in vitro anticancer activity to in vitro toxicity on human bone marrow as follows: cisplatin 0.1 micrograms/ml, doxorubicin 0.01 micrograms/ml, etoposide 0.1 micrograms/ml, mitomycin-C 0.005 micrograms/ml, vindesine 0.01 micrograms/ml. At these concentrations, clinically sensitive tumor types showed inhibition of colony formation in 99 of 240 cases (41%), whereas 11% (19/176) of clinically resistant tumors were responsive. The relevant dose levels used equal between 0.3% and 4.0% of the achievable peak plasma concentrations in man. The predictive value of the clonogenic assay was determined by treatment of the same tumors in vitro and in vivo in tumor-bearing nude mice. In 174/220 comparisons (79%), in vitro data predicted correctly for the in vivo sensitivity of the xenografted malignancies.(ABSTRACT TRUNCATED AT 250 WORDS)     

Anticancer drug activity in human bladder tumor cell lines

A panel of ten human bladder tumor cell lines were tested for drug sensitivity to ten standard or investigational anticancer drugs using a tumor colony assay. The activity of these anticancer agents in vitro was then compared with the clinical activity of these agents in bladder cancer. Drug activity was found in only five of the ten cell lines. In only 9 of 100 drug assays was the inhibition of colony growth lower than 30% of the controls. The activity of the more active anticancer drugs in bladder cancer (i.e., methotrexate and cisplatin) was not predicted using the tumor colony assay. Overall, the low level of activity of most anticancer drugs tested paralleled the clinical experience of drug resistance found in human bladder cancer.     

Combined recombinant human interferon alpha 2 and cytotoxic agents studied in a clonogenic assay

A human tumor clonogenic assay has been used to test the antiproliferative effect of recombinant human leukocyte interferon alpha 2 alone and in combination with each of 8 cytotoxic agents. Cell lines derived from 6 human tumors and primary tumor cells from 13 patients have been used in these clonogenic assay studies. Results show that interferon as a single agent causes insignificant reduction in tumor cell colony survival if the short-term 1-hr cell exposure method is used; only high concentrations of interferon used in continuous cell exposure in the clonogenic assay can demonstrate a reduction in colony survival to below 50% of control values. Combinations of interferon with either doxorubicin or cisplatin frequently show additive and occasionally synergistic antiproliferative effects on tumor cell colony formation. Variations in drug concentrations and sequencing of drugs have been tested, showing that optimal antiproliferative effects of combined interferon and doxorubicin are realized when maximal concentrations of interferon and prolonged cell exposure time of both interferon and doxorubicin are employed. Combinations of interferon and doxorubicin tested in the clonogenic assay demonstrate cytotoxicity superior to that of either agent tested alone.     

Thermochemosensitivity: augmentation by hyperthermia of cytotoxicity of anticancer drugs against human colorectal cancers, measured by the human tumor clonogenic assay

Thermochemosensitivity was examined in vitro by a human tumor clonogenic assay (HTCA) using specimens obtained surgically from 43 patients with colorectal cancer. We found that the percentages of patients whose cells showed higher sensitivity (greater than 70% inhibition of colony formation) to hyperthermia alone were 29.6 and 55.6% at 42 and 43 degrees C each for 1 h, respectively. Similarly, percentages of patients whose cells were sensitive to drugs alone were 13.8, 24.1 and 48.3%, for cis-diamminedichloroplatinum(II), 5-fluorouracil and mitomycin C, respectively. These sensitivities were augmented when hyperthermia was combined with administration of anticancer agents. Even in tumor cells that were insensitive to anticancer drugs alone or to hyperthermia alone, sensitivity was enhanced when administration of drugs was combined with hyperthermia. Our results demonstrate that HTCA of tumor cells, obtained from suitable patients, may be useful as a predictive test for application of thermotherapy and for individualization of chemotherapy.     

In vitro chemosensitivity of urological malignancies evaluated by colony-forming assay using soft agar

Over the past year, we have attempted to grow 132 different human tumor samples (78 from urological and 54 from non urological tumors) using a soft agar colony formation assay similar to that originally described by Salmon and colleagues. Formation of colonies in vitro occurred in 44 of 78 primary urological tumors (56%), including 63% (12/19) of renal cancers, 61% (25/41) of uroepithelial cancers and 42% (5/12) of testicular cancers. The effects of in vitro chemosensitivity were analysed using the inhibition of colony growth (more than 70%) at two different cut-off doses which were equilibrated with the achievable AUC doses for the higher cut-off point and to one-tenth of AUC for the lower cut-off point. Five of 13 (38%) drugs showed an effective rate between 10% and 38% for the lower cut-off point in vitro. On the other hand, eleven of 13 (85%) drugs. Showed an effective rate between 20% and 67% for the higher cut-off point in vitro. In the tested uroepithelial cancers, none of seven for the lower cut-off and three of seven for the higher cut-off were demonstrated to be sensitive cases from the dose corresponding colony inhibition curve for cisplatinum. Nine of the renal cancers were demonstrated for the dose corresponding colony inhibition curve for Interferon. To predict clinical correlation, 16 patients (20 drugs) were treated with identical drugs which were estimated from in vitro chemosensitivity testing. The predictability results were 75%-100% true positive and 85%-100% true negative, with 87% overall predictability. This assay can therefore be used to study differences of biological character including drug sensitivity.     

Chemosensitivity testing of human colorectal carcinoma cell lines using a tetrazolium-based colorimetric assay

The in vitro chemosensitivity of 11 human colorectal cell lines to seven chemotherapeutic agents was determined using a semiautomated tetrazolium-based colorimetric assay (MTT assay). Four of the cell lines were from primary tumors and seven from metastases. Eight lines were from patients with no prior chemotherapy. From assay results, we predict 5-fluorouracil (5-FU) to be the sole active agent of the seven tested. This is based on two observations: the range of drug concentrations which produced 50% inhibition of cell growth was greatest with 5-FU (388-fold versus 5- to 30-fold with the other six agents); and the area under the curve (AUC) which produced 50% growth inhibition was within a clinically achievable range only for 5-FU. Since the assay AUC of 5-FU at 50% inhibition was in a clinically achievable range for only two of the 11 cell lines, we performed a multivariate analysis to explore parameters which predict 5-FU sensitivity. In the best fitting model, sensitivity was positively correlated with cloning efficiency in media and with cell surface TAG-72 (a tumor-associated glycoprotein found on epithelial tumors of ovary, lung, colon, and breast origin) expression. If validated with an in vivo test such as the nude mouse model, the MTT assay could be very useful in new drug screening for colorectal carcinoma, for examining combination chemotherapy for synergy, for exploring strategies for biochemical modulation, and perhaps in individualizing therapy when cell lines can be established from a patient.     

p73基因过量表达对人肺癌细胞A549凋亡及其化疗敏感性的作用

背景与目的：部分非小细胞肺癌（non-small cell lung cancer,NSCLC）表达野生型p53基因（wild-type p53,wt-p53）,因此在基因治疗中克服这些NSCLC对wt-p53的抵抗机制就非常重要。P53基因家族的新成员p73是p53的同源体,本研究旨在探讨对wt-p53基因治疗抵抗的人肺腺癌细胞A549在外源p73基因转染或联用化疗药后凋亡程度和对化疗药物敏感性的变化。方法：将真核表达重组质粒pcDNA3-HA-p53或pcDNA3-HA-p73α转染入A549细胞,G418筛选,Western blot检测P53或P73α的表达。MTT法分析转染细胞对顺铂和阿霉素的敏感性,用流式细胞术、TUNEL法和DNA片段法分析化疗药作用下转染细胞的凋亡变化,克隆形成实验观察细胞生物学性状的改变。结果：转染p53或p73α基因的A549细胞可以稳定高表达p53或p73α蛋白。转染p73α的A549细胞在原本没有明显抑制和杀伤作用的药物浓度（6．25μmol／L的顺铂或0．25μmol／L的阿霉素）作用下生长明显受到抑制。顺铂的IC50值从22．65μmol／L降至3．75μmol／L,阿霉素的IC50值从4．20μmol／L降至0．06μmol／L。p73能使A549细胞受顺铂和阿霉素诱导的细胞凋亡增加。而p53没有明显作用。流式细胞术显示p73α基因转染后,顺铂诱导的细胞凋亡率从10．6％升高到36．8％（P〈0．01）,阿霉素诱导的细胞凋亡率从13．0％升高到41．1％（P〈0．01）。克隆形成实验显示．p73α基因转染能明显降低顺铂和阿霉素作用后A549细胞的克隆形成数（P〈0．01）。对顺铂和阿霉素的化疗增效倍数分别为2．0和2．4倍。结论：外源性p73基因的导入增加了wt-p53型A549细胞对顺铂和阿霉素等化疗药的敏感性。该作用可能与p73基因能不依赖于p53基因诱导细胞凋亡有关。p73基因可用于治疗wt-p53不能发挥作用的恶性肿瘤。     

ＡＴＰ生物荧光体外药敏检测在中晚期结直肠癌化疗药物筛选中的应用

目的　探讨ＡＴＰ生物荧光体外药敏检测法（ＡＴＰ-ＴＣＡ）的特点及其在中晚期结直肠癌患者化疗方案中的指导价值。方法　应用ＡＴＰ-ＴＣＡ体外检测５９例结直肠癌细胞对常用抗癌药物的敏感性。结果　ＡＴＰ-ＴＣＡ法对结直肠癌标本的可评估率为９６.６１％。５７例结直肠癌细胞对４组联合化疗药物氟尿嘧啶＋丝裂霉素、氟尿嘧啶＋奥沙利铂、氟尿嘧啶＋伊立替康、氟尿嘧啶＋奥沙利铂＋伊立替康相比氟尿嘧啶、丝裂霉素、伊立替康、奥沙利铂４种单药的敏感度有高度显著性差异（Ｐ＝０.０００６）。应用ＡＴＰ-ＴＣＡ检测结果指导中晚期结直肠癌患者化疗,临床近期有效率为５９.６５％（３４／５７）,总预测准确率为６３.１６％（３６／５７）,阳性符合率为６１.８２％（３４／５５）,阴性符合率为１００％（２／２）。结论　ＡＴＰ-ＴＣＡ能有效检测化疗药物的敏感性,对指导中晚期结直肠癌患者化疗有重要的临床意义。     

A Southwest Oncology Group study on the use of a human tumor cloning assay for predicting response in patients with ovarian cancer

A total of 211 patients with epithelial ovarian cancer (168 with tumors refractory to prior chemotherapy and 43 with no prior chemotherapy) from 33 different Southwest Oncology Group institutions had their tumors sampled and specimens shipped to two central laboratories for drug-sensitivity testing in a human tumor cloning assay. The 168 patients with a prior history of chemotherapy failure (median of four prior chemotherapeutic agents) were treated with the most effective agent(s) found in the cloning assay (23 patients), and those patients whose tumors did not form colonies in vitro or did not manifest any sensitivity to agent(s) were treated with a clinician's choice of agent(s) (101 patients). The remaining 44 of the 168 patients were not treated with chemotherapy because of deteriorating performance status or early death. The complete and partial response rate in patients treated according to assay results was 28% versus 11% for the patients treated according to clinician's choice (P = 0.03). There was no statistically significant difference in survival between the two options (6.25 versus 7 months, respectively). The 43 patients with no history of prior chemotherapy were all treated with standard combination chemotherapy, and their clinical response was compared with their in vitro sensitivity to the same agents. Overall there was a 100% true-positive rate and 100% true-negative rate for the seven evaluable patients. From these data the authors conclude that use of the human tumor cloning assay may increase the response rate but not the survival for selected patients with advanced chemotherapy-refractory ovarian cancer. The study is weakened, however, by the many steps of patient selection necessitated by inadequate tumor colony formation in vitro and the inability to treat all patients (because of early death or a rapid decline in performance status). The assay does appear to be worthy of additional study for predicting response to combination chemotherapy in patients without a prior history of chemotherapy. Finally the use of central chemosensitivity testing laboratories is feasible for testing in vitro predictive assays in a cooperative group setting.     

Effects of suramin on in vitro growth of fresh human tumors.

BACKGROUND: Suramin is a polysulfonated urea recently tested in clinical trials as an anticancer agent. PURPOSE: To define tumor types for further clinical testing of suramin, we assessed the in vitro activity of suramin against fresh human tumor specimens. METHODS: Inhibition of tumor colony formation (human tumor clonogenic assay [HTCA] method) and inhibition of tritiated thymidine incorporation (TTI method) were used as indicators of drug sensitivity. RESULTS: With the use of the HTCA method, 80% or more of carcinomas of the colon, endometrium, kidney, lung (non-small-cell), and ovary as well as malignant melanoma and mesothelioma were sensitive to 200 micrograms/mL of suramin by continuous exposure. Suramin's antitumor activity was dose dependent, and it was less effective when tested at concentrations of 50 micrograms/mL or less. With the TTI method, the more slowly growing tumors (breast cancer, colon cancer, multiple myeloma, non-Hodgkin's lymphoma, prostate cancer, and sarcoma) appeared to be less sensitive to suramin. However, when the two assay methods were directly compared in melanoma and ovarian cancer specimens, individual tumors were generally more sensitive to suramin (greater inhibition relative to control) using the HTCA method, whereas the TTI method appeared to underestimate suramin's antitumor activity. There was no significant difference in the activity of suramin when tested in the presence of 10% versus 50% serum. CONCLUSION: These results provide an experimental basis for clinical evaluations of suramin therapy in patients with colon, endometrial, kidney, non-small-cell lung, and ovarian cancers as well as malignant melanoma and mesothelioma.     

Chemosensitivity of adriamycin, mitomycin C, and bleomycin with in vitro colony assay

In vitro colony assay was used to examine the sensitivities of adriamycin (ADM), mitomycin C (MMC), and bleomycin (BLM), and the effects of a prior chemotherapy on in vitro chemosensitivities of the drugs. One hundred forty-seven tumor specimens from 106 patients with various malignancies were placed in culture. Seventy tumors (56.0%) from 125 evaluable specimens which had histologically or cytologically confirmed to be malignant, and were plated at 5 x 10(5) cells/plate formed more than 30 colonies/plate enough for sensitivity assays. The results of in vitro sensitivities of the drugs were nearly similar to those reported in the clinical trials. The in vitro antitumor activity of BLM was noted in 3 (21.4%) of 14 patients with ovarian cancer, therefore the results suggest that the assay is useful to select effective drugs against tumors in which phase II study has never been performed. In vitro sensitivity of BLM was markedly reduced by a prior chemotherapy containing the same drug (p less than 0.05), and prior therapeutic ADM exposure also reduced in vitro activity of the drug, whereas there was no significant influence on in vitro activity of MMC. These results indicate the usefulness of the in vitro colony assay for predicting clinical antitumor activities of ADM, MMC, and BLM.