Identification of genes overexpressed in tumors through preferential expression screening in trophoblasts.

Early trophoblastic cells share several features with neoplastic cells. Based on that observation, we attempted to identify genes overexpressed in tumors by analyzing genes preferentially expressed in trophoblasts. A subtracted library enriched in complementary DNA from early cytotrophoblasts was constructed, and the expression level of selected recombinants was analyzed on a large panel of normal and tumor tissues. The library was prepared using a polymerase chain reaction-based complementary DNA subtraction method with 6-week amenorrhea cytotrophoblast endoplasmic reticulum-bound RNA as target, and a mixture of complementary DNA prepared from terminal placenta and activated T-lymphocytes as driver. Two rounds of screening were performed to isolate clones preferentially expressed in early placenta. From a total number of recombinant clones estimated at 32,000 in the subtracted library, 594 inserts were analyzed by Southern blot and 21 sequences were isolated as corresponding to genes highly expressed in early placenta. Eleven encoded known molecules, such as carcinoembryonic antigen, human chorionic gonadotropin, and mitochondrial rRNAs. Ten sequences represented novel genes. Northern blot analysis confirmed that most of these genes were preferentially expressed in early trophoblast in comparison to terminal placenta. Three clones that gave detectable hybridization signals on total RNA were extensively studied and were found to be overexpressed in various tumors. Two of these clones, designated B9 and E4, were later identified as corresponding to genes coding for the putative ribosomal protein S18 and the bifunctional enzyme ADE2H1 involved in purine biosynthesis, respectively. Expression of the third clone, E9, was increased up to 10-fold in breast cancer tissues in comparison with normal counterparts. Present results confirm that many genes expressed in the trophoblast are overexpressed in malignant cells. This approach could provide a general targeted method for the identification of genes overexpressed in various neoplastic cell types.

Tumor-infiltrating CD3- NK cells are more effective than CD3+ T cells in killing autologous melanoma cells.

We have studied the phenotype and functional activity of tumor-infiltrating lymphocytes (TIL) derived from eight human melanomas cultured for up to 60 d in the presence of recombinant IL-2. In the early period of the cultures, TIL were predominantly T cells of CD8+ phenotype and contained 10-30% of CD3- cells. Four of the five early TIL cultures tested in a cytotoxicity assay displayed a degree of MHC-unrestricted lysis on a series of autologous and allogenic melanoma cell lines as well as the K562 natural killer-sensitive target. With longer periods of time in culture, all TIL lines showed a decrease in lytic activity that was associated with the loss of CD3- cells. Thus, most of the killing of short-term TIL cultures appeared to be mediated by CD3- natural killer cells, whereas CD3+ T cells were found to be weak anti-tumor effectors. Even though the CD3+ T cells were not cytotoxic on K562 targets, their lytic activity (even weak) against melanoma cells appeared to be non-MHC restricted, and was blocked by anti-CD3 antibodies. In addition, cytotoxicity of the CD3+ TIL cultures was compared to that of a CD3-/NKH1+ cell line purified from peripheral blood. It was found that natural killer cells were much more potent than CD3+ TIL on the melanoma cell lines tested.

Prognostic value of neural invasion in rectal carcinoma: a multivariate analysis on 339 patients with curative resection.

To determine whether neural invasion or other clinico-pathological factors are prognostic, we performed a retrospective study on 339 rectal carcinomas. The overall 5-year survival was 62%. In the multivariate analysis, age over 60 years, a distance from the anal verge of less than 6 cm, the number of positive lymph nodes, neural invasion and tumour penetration were found to be prognostic. A scoring system identified five prognostic groups of patients. Neural invasion is an independent prognostic factor in our scoring system and it is suggested that this parameter should be taken into consideration for postsurgical treatment.

A comparison of early effects with two dose rates in brachytherapy of cervix carcinoma in a prospective randomised trial.

This phase III randomised trial examined the early effects of two low dose rates (0.38 and 0.73 Gy/h) in brachytherapy of stage I and IIp cervical cancer patients. A total of 204 patients were included between January 1985 and September 1988. Since the main analysis of this paper concerned surgical difficulties, only the 155 patients (76%) on whom surgery was performed at the Institut Gustave-Roussy were retained in this analysis. Treatment consisted of uterovaginal 137Cs irradiation followed by immediate or deferred surgery. The two groups were similar for pretreatment characteristics except for endocervix involvement. Their brachytherapy parameters were also similar (60 Gy pear dimensions, doses to critical organs, total kerma, etc.). The factors with a poor prognosis were, for surgical difficulties, older age, stage II and a small irradiated pear volume; for difficulties with haemostasis, immediate surgery, stage II and previous surgery; and for difficulties in dissection, lymph node involvement. The dose rate significantly influenced surgical difficulties for the stage IIp patients operated on by deferred surgery. Those treated with the higher dose rate showed a 2-fold increase in surgical difficulties compared to those irradiated at the lower dose rate (P = 0.03). The independent prognostic factors for sterilisation of the surgical specimen were small tumour size and absence of lymph node involvement. An inverse dose rate effect was observed for medium size tumours, with significantly more sterilisations observed in stage IIp patients in the lower dose rate group (P < 0.01).

Whole abdominal irradiation following chemotherapy in patients with minimal residual disease after second look surgery in ovarian carcinoma.

From January 1981 through December 1985, 65 patients with epithelial carcinoma of the ovary were treated with the following protocol: surgery, combination chemotherapy, second-look surgery documenting tumor less than or equal to 2 cm, and whole abdominal irradiation. Chemotherapy consisted of a combination of cyclophosphamide, adriamycin, and cisplatinum in 89% of the patients. The median number of cycles was eleven. Second-look surgery documented no residual tumor in 23 patients, microscopic disease in three patients, and macroscopic disease less than or equal to 2 cm in 39 patients. Whole abdominal irradiation was given with an open field technique up to 20 Gy without renal or hepatic shield. A pelvic boost of 15-30 Gy was subsequently added in 17 patients with macroscopic disease in the pelvis at the time of second-look surgery. Fifteen patients received complementary chemotherapy mostly hexamethylmelamine. All but two patients completed whole abdominal irradiation: one refused further radiotherapy after 3 Gy and one developed disease progression with bowel obstruction after 1 Gy. The median follow-up was 69 months. The 3-year and 6-year no evidence of disease survival rates were 60% (95% CI: 48-71) and 33% (95% CI: 21-46), respectively. The 3-year and 6-year recurrence rates were 33% (95% CI: 22-45) and 54% (95% CI: 40-67), respectively. The 3-year and 6-year metastasis rates were 22% (95% CI: 13-34) and 43% (95% CI: 30-58), respectively. A multivariate analysis showed that residual disease after second-look surgery was the only significant prognostic factor with a relative risk of death or local or distant failure of 4.2 (95% CI: 1.9-9.5, p less than 10(-4)). Two patients developed mean-term gastrointestinal complications (small bowel obstructions requiring surgery). Survival remains poor with high level of failure even with aggressive multimodal treatment.

Prospective comparison of ultrasound and computed tomography in the evaluation of the size of the uterus: can these methods be used for intracavitary treatment planning of carcinoma of the uterus?

A prospective evaluation of computed tomography and ultrasound was performed on 34 patients with Stage I and II endometrial carcinoma. All patients underwent immediate surgery following intracavitary treatment directed to vaginal mucosa. Pathologic measurements of the uterus were compared to those obtained by imaging technologies. The results of the study suggest that all but height measurements were rather accurately determined by both ultrasound and CT scan. However ultrasound was significantly better in determining the size of the cervix. Therefore ultrasound measurements could be used routinely for intracavitary treatment planning in endocervical carcinoma and endometrial carcinoma.

Combined radiotherapy and surgery: local control and complications in early carcinoma of the uterine cervix--the Villejuif experience, 1975-1984.

From January 1975 to December 1984, 441 patients were treated by combined radiotherapy and surgery at the Institut Gustave Roussy (IGR) for Stage IB (288) and II (proximal) (103) carcinoma of the uterine cervix. Standard treatment consisted of pre-operative utero-vaginal brachytherapy (60 Gy) using a mould technique followed by a colpo-hysterectomy and external iliac lymphadenectomy. Overall 5 year actuarial survival for the whole population was 87% and disease-free survival 85%. Loco-regional relapse occurred in 23 patients (5%). Of these, 12 were central pelvic failures, 8 regional failures and 3 combined central and regional failures. There were 36 systemic relapses (8%) of which 12 relapsed concurrently in the pelvis. Five year actuarial pelvic disease-free, disease-free and overall survival was 87, 85 and 87%, respectively, for the whole population. 340 patients developed one or more complications [Grade 1: 198/441 (44%), Grade 2: 121/441 (27%) and Grade 3 or 4: 21/441 (4.7%)]. Five year actuarial survival for the whole population was poorer for histologically node positive than for node negative (89 vs. 55%, p less than 0.0001). Pre-operative brachytherapy followed by surgery can provide good local control with acceptable morbidity in early cervical cancer.

Malacoplakia of the gallbladder.

A case of malacoplakia of the gallbladder is described. The cytoplasm of histiocytes in the gallbladder wall was filled with granules positive for periodic acid-Schiff, von Kossa's, and Perls' stains, which is highly suggestive of malacoplakia. Both local inflammation and recent neoplasia could have played a role in the histogenesis of the malacoplakia.

Gastric carcinoma with argyrophilic cells: light microscopic, electron microscopic, and immunochemical study.

An immunochemical study of a gastric adenocarcinoma with argyrophilic cells showed two areas of tumor that react differently with the usual histochemical reagents as well as with immune sera against gastrin and mucoprotein associated with antigens. Ninety per cent of the tumor cells were PAS positive and contained M2 antigen, and some also contained M1 antigen. About 30 per cent of the M2-positive cells stained strongly with an antigastrin serum as well as with the argyrophilic reagents. The remaining 10 per cent of tumor cells were signet-ring cells located in several clumps in the tumor. These cells were positive for both PAS and alcian blue and contained intestinal M3 antigen. Forty-five per cent of them also contained M1 gastric antigens. Carcinoembryonic antigen (CEA) was found in the cytoplasm of each tumor cell. The presence of CEA and M1 antigen together indicates a fetal pattern, suggesting that the cells originate from very immature gastrointestinal stem cells.

Relationship between the melanin content of a human melanoma cell line and its radiosensitivity and uptake of pimonidazole.

The intra-cellular uptake of the weakly basic radiosensitiser pimonidazole (PIMO) was determined as a function of the pigmentation of Na11+ human melanotic melanoma cells in vitro. Two experimental conditions were considered: exponentially growing cells (Exp.) and plateau-phase cells (PI.). The melanin content of Na11+ cells ranged from 500 micrograms/g cell weight in exponentially growing cells to 6000 micrograms/g in heavily pigmented plateau-phase cells. Cells were exposed to PIMO (medium dose, 0.2 mmol/dm3; 58.2 micrograms/ml). The intra-cellular concentration ranged from 163 micrograms/g in Exp. to 900 micrograms/g in pigmented Pl.; the latter being equivalent to an intra- to extracellular concentration ratio (Ci/Ce) of 17. However, this increase in the cellular uptake of PIMO was not accompanied by an increase in radiosensitising efficiency. In comparison, the Ci/Ce for etanidazole (ETA), a radiosensitiser that is uncharged at physiological pH, remained approximately constant at 1 for all values of melanin contents. Treatment of Na11+ tumours in vivo with [3H]-PIMO resulted in a tumour:blood ratio of about 3 at 30-60 min after administration. However, at 24 h a grain count of label derived from [3H]-PIMO showed that picnotic areas of tumours contained levels that were some 40 times greater than the background value. This high level of label was coincident with areas of highest apparent melanin content. In conclusion, PIMO accumulates in very heavily pigmented melanoma cells present in necrotic zones with picnosis. As these cells are probably non-clonogenic, PIMO is not suitable for use in melanoma radiotherapy.

Comparison between familial and nonfamilial melanoma in France.

BACKGROUND AND DESIGN: Five percent to 10% of cutaneous malignant melanomas (CMMs) occur in a familial setting. Clinical, epidemiologic, and genetic studies of familial CMM in different regions of the world have led to various results. To assess the characteristics of familial CMM in France, the clinical, histologic, and epidemiologic characteristics of 295 patients with CMM were recorded, and a comprehensive familial investigation was performed for each case. Patients with a family history of CMM were compared with nonfamilial cases. RESULTS: Cutaneous malignant melanoma occurred as a familial cancer in 22 (8%) of 295 patients. Among the multiple variables studied, those significantly associated with the familial occurrence of CMM were red hair, inability to tan, and presence of clinically atypical moles. When these variables were considered together in a multivariate analysis, only the association with red hair (P = .001) and atypical moles (P < .05) remained significant. In addition, the patients with familial melanoma exhibited the following tendencies: a younger age at diagnosis, a higher number of moles, and the development of multiple primary melanomas, but these results did not reach statistical significance. Factors relating to UV light exposure, histologic features of CMM, course of the disease, and occurrence of nonmelanoma cancers showed a similar distribution between familial and nonfamilial cases. CONCLUSION: A familial investigation should be performed for each patient with CMM in France, particularly when he or she exhibits phenotypic risk factors for CMM such as red hair and atypical moles.

Secondary cytoreductive surgery in ovarian cancer.

During the 8-year period from 1976 to 1984, 202 patients with a primary ovarian cancer underwent a second-look laparotomy at the Institut Gustave-Roussy (Villejuif). One hundred and nine patients had a macroscopic tumour, in 77 of which there was clinical evidence of disease before the laparotomy. Fifty-seven patients underwent an optimal resection of the tumour (largest residual tumour less than 2 cm) and 52 underwent non-optimal cytoreductive surgery or isolated biopsies. In 22 cases the optimal resection necessitated a bowel resection. Survival curves suggest: (1) that the removal of macroscopic residual disease does not improve life expectancy except in the cases of optimal resection without bowel resection. (2) When there is evidence of disease before the second-look operation the prognosis remains the same whatever the surgery performed.

Melanoma arising de novo in childhood: experience of the Gustave-Roussy Institute.

Between January 1956 and December 1990, 17 patients younger than 17 years with available pathological screens of de novo cutaneous melanoma, and with no other risk factors (xeroderma pigmentosum, giant congenital naevi, congenital melanoma or a proven family history of dysplastic naevus syndrome) were seen at the Gustave-Roussy Institute. The median age was 9 years and 9 months (range 2 years and 3 months-16 years and 9 months). The primary disease was located in the lower extremities in 10 cases, the trunk in five cases, and the upper extremities or head and neck in one case. The disease was localized for 10 patients at presentation (stage I), six had proven nodal metastasis (stage II) and one patient had nodal and breast metastases. The median thickness of the primary lesion was 2.89 mm (range 0.64-10). Five tumours were at level III on Clark's index, eight at level IV and four at level V. Six cases were classified as superficial spreading, two as unclassified radial growth, three nodular, three with Spitzoid cells, and three were unclassified. Two patients presented local recurrence with an initial unclassified melanoma, with a thickness greater than 2.5 mm. At a median follow-up time of 7 years, two patients had died from recurrent disease, and one patient had died from a second malignancy.

Tumour oxygenation, radiosensitivity, and necrosis before and/or after nicotinamide, carbogen and perflubron emulsion administration.

Hypoxia is one of the factors involved in tumour resistance to radiotherapy. One way to improve tumour oxygenation is to use oxygen carriers such as perflubron emulsion plus carbogen or vasoactive drugs such as nicotinamide. The perflubron emulsion and carbogen act mainly on hypoxia caused by limited diffusion of oxygen; nicotinamide acts mainly on acute hypoxia. The aim was to correlate radiosensitivity and pO2 measurements (computerized pO2 histograph) after nicotinamide, perflubron emulsion and carbogen administration, and to determine the role of necrosis in this correlation. Two human tumour xenografts (HRT18, Na11 +) and one rodent tumour (EMT6) were used. Clonogenic assays and pO2 measurements were performed under similar conditions. The radiosensitization and oxygenation levels increased with all treatments. The maximal effects were found with the combination of nicotinamide (1 g/kg), perflubron emulsion and carbogen. A correlation between the radiosensitization and the pO2 measurements was found for the three cell lines with a cut-off point of 10 mmHg. The presence of necrosis could explain the low pO2 (< 2 mmHg) found even when complete radiosensitization was observed.

International pathologists congruence survey on quantitation of malignant melanoma.

A congruence survey of pathologists in Brisbane, Adelaide, Oslo and Paris on classifying and grading histological features such as dermal invasion, cross-sectional profile, mitotic activity and lymphocytic infiltrate was done on 147 cutaneous melanomas. The overall agreement was about 70%, about one pathologist in three or four disagreeing on each feature. Taking into acount also the considerable number of slides in which agreement was less, the results are considered unsatisfactory. Agreement was highest in cross-sectional profile and least in level of invasion. It is evident that the gradings are subjective and indicate the need for detailed histological description in making the criteria and the necessity of collaboration between pathologists to study slides and clarify problems. Experience is important and the slides must be of high technical excellence. Comparisons of results between different pathologists on the histological quantitations of malignant melanoma should take note of the degree of congruence achievable.

Glutathione and cysteine levels in human tumour biopsies.

There is evidence that some human tumours could be treated with a combination of buthionine sulfoximine and hypoxic cell sensitizers. However, clinical application of this technique requires a prior knowledge of the level of non-protein bound sulfhydryl (NPSH) compounds in these tumours. The present study provides data on the levels of glutathione (GSH) and cysteine (CYS) in human tumour biopsies from the cervix and from the head and neck. The NPSH compounds were measured by high performance liquid chromatography. The median GSH values were 20.5 nmol/mg protein (cervix) and 23 nmol/mg protein (head and neck) while the median CYS values were 4.4 (cervix) and 4.2 nmol/mg protein (head and neck). The values varied widely from one patient to another.

Fine needle aspiration of the liver and pancreas with ultrasound guidance.

The experience of the Institut Gustave-Roussy in the diagnosis of hepatic and pancreatic lesions by fine needle aspiration (FNA) is reported. Totals of 116 consecutive percutaneous ultrasound-guided FNAs of the liver and 27 of the pancreas were performed without complication in patients with ultrasonically suspected neoplastic lesions. In 12 cases, the material was not suitable for diagnosis. In the liver, 97 cases were correctly diagnosed and confirmed by follow-up. Immunohistologic studies were helpful in distinguishing primary liver tumors from other malignancies. One false-positive result was reported. In the pancreas, malignancy was detected in 17 cases. Cytology alone provided the correct tumor diagnosis in 15 cases: 10 primary carcinomas, 2 endocrine tumors and 3 metastases. The sensitivities of FNA in this study were 87.6% for the liver and 85% for the pancreas, similar to those reported in other series.

[Anatomical extension of invasive carcinoma of the uterine cervix (author's transl)]. / Extension anatomique des carcinomes infiltrants du col utérin.

Invasive epithelioma of the uterine cervix spreads in two different ways. Locally, tumour involves progressively upon neighbouring structures (vagina parametrium, uterine corpus, bladder, rectum). Regionally, lymphatic dissemination occur preociously moving towards the nodes of the pelvic wall. Local extension can be appreciated by clinical examination which determines the clinical staging of cervical cancer. It does not take sufficiently into account the volume of the tumour which is a principal factor in lymphatic dissemination. Nodal metastases essentially localise in the external iliac chain (obturateur group), from there spreading to the common iliac or latero aortic nodes which may, under exceptionnal circumstances be involved primarily. The incidence of latero-aortic metastases has recently been high-lighted by pre-treatment staging laparotomy. Besides lymphatic metastases, systematic metastases via a venous spread are rare. They indicate a late diagnosis or therapy failure with pelvic relapse. The therapeutic effort in cancer of the uterine cervix, at the present time, ought to be found on loco-regional treatment using either radiotherapy or surgery.

[Role of electrocoagulation in treatment of cancers of the rectum]. / La place de l'électrocoagulation dans le traitement des cancers du rectum

The authors describe the technics of a wide local electrocoagulation which was used in the treatment of 42 malignant tumors of the rectum: 27 adenocarcinomas and 15 malignant villous tumors. This not disabling technic, avoiding colostomy, is generally harmless even in old and poor risk patients. The mortality rate was 7 p. 100 in the adenocarcinomas series, nul in the malignant villous tumor series. The five year survival rate was 66 p. 100 in the first series and 80 p. 100 in the second. One third of the patients bearing adenocarcinomas developed recurrences, among which some were treated either by large surgery, iterative electrocoagulation, or radiotherapy after colostomy. The indications depend on the size and the site of the tumor. Electrocoagulation is found possible only for T1 or T2 tumors of the posterior and lateral walls of the rectum, provided that they are located at less than 10 cm from anus and above the sphincter.

[Presence of free retinol receptor (cRBP) and retinoic acid receptor (cRABP) in human skin tumors]. / Présence du récepteur libre au rétinol (cRBP) et du récepteur à l'acide rétinoïque (cRABP) dans les tumeurs cutanées humaines.

The concentrations of cellular retinoic acid binding protein (cRABP) and free cellular retinol binding protein (cRBP) were determined by ultracentrifugation in sucrose gradient in the cytosol of 41 human skin tumours (14 melanomas, 19 basal cell carcinomas, 8 squamous cell carcinomas). cRBP was found respectively in 36%, 42% and 37% of the studied samples. On the contrary, cRABP was more frequently found in carcinomas (89% in basal cell carcinomas and 100% in squamous cell carcinomas) than in melanomas (21%) (p less than 0.001). These results are discussed according to the different embryologic origin of carcinomas and melanomas. Furthermore, the better efficiency of synthetic retinoids in carcinomas than in melanomas should be explained by a different way of action in these 2 kinds of tumours.