Chromosomal anomalies in children treated for cancer.

Cytogenetic studies were carried out in peripheral blood lymphocytes of 31 children with cancer (12 retinoblastomas, 9 non-Hodgkin's lymphomas, 7 neuroblastomas and 3 Wilms' tumors) with the purpose of investigating the prevalence and persistence of chromosomal aberrations before onset of antineoplastic treatment and at different intervals thereafter. The number of treated patients with chromosomal anomalies was significantly higher (p less than 0.01) and so was the percent of cells with aberrations, as compared to untreated patients or healthy controls. The most frequent aberrations were of chromatid type. One patient had a number five trisomy in the third posttreatment study and another presented abnormal aneuploid cells in the third and fourth posttreatment studies. Congenital chromosomal anomalies were not observed, nor anomalies described for some of these tumors. There was no regularity in the effected chromosomes. We conclude that the observed chromosomal aberrations were due to treatment. We consider a prolonged and stringent cytogenetic follow up of such patients necessary to detect the induced aberrations.

Antitumor activity of four polymer bound trans-1,2-diaminocyclohexaneplatinum(II)-4-carboxyphtalate complexes tested in different model systems.

Four polymer bound Pt-complexes have been tested in in vivo and in in vitro systems. No substantial difference in effectivity against P388 leukemia in vivo was found when free trans-1,2-diaminocyclohexaneplatinum(II)-4-carboxyphtalate (TMA) was compared with the polymer bound complexes. The compound with the highest ID50 value in soft agar assay exhibited low effectivity in in vivo testing. Polymer bound Pt-complexes with faster release of the active molecule exhibited in in vivo and in soft agar assay slightly lower activity, when compared with suspension culture test system. Cross resistance of polymer bound complexes was investigated on three cell lines with induced drug resistance against different Pt-complexes. Cross resistance was found between TMA (free and polymer bound) and trans-1,2-diaminocyclohexaneplatinum(II)citrate (PEX) as well as trans-1,2-diaminocyclohexaneplatinum(II)glucarate (PTU) but there was no cross resistance between TMA and cis-diamminedichloroplatinum(II) (cis-DDP).

The "in vivo" effect of protein A on tumor growth.

The antitumor effect of two purified preparations of protein A obtained from Staphylococcus aureus strain Cowan 1 or the mutant strain A676 was investigated. This study using different doses of SpA (5 or 10 mg/animal) i.p. administered was performed in lymphosarcoma bearing rats at certain intervals of time after tumor transplantation. The treatment with SpA deriving from A676 strain which was started at the same time with the tumor transplantation led to a significant increase in the latency period of tumor growth and the prolongation of survival time of tumor bearing rats. Attempts to use a higher dose of SpA and also a smaller tumor inoculum did not improve the results with regard to inhibition of lymphosarcoma growth in inbred R rats.

Concanavalin A-induced agglutination of Ehrlich ascites carcinoma cells during growth of the tumor in mice.

ConA-induced agglutination of Ehrlich ascites carcinoma (EAC) cells during development of the tumor in mice has been studied in vitro. EAC cells were incubated with different concentrations of ConA at 37 degrees C for various intervals of time and the cell aggregates of different sizes were scored under microscope. The older EAC cells (20-day-old) showed a decrease in cytoagglutination with respect to large aggregates formation compared to the younger (10-day-old) ones. The small aggregates formed by the 5-day-old EAC cells are dissociated by competition with alpha-methyl-D-mannoside but the large aggregates are not affected by the sugar treatment. The observed differences in the agglutination patterns are discussed in view of the influence of the lipid content of EAC cells and the ascites fluid on ConA-induced cytoagglutination.

Influence of ascorbic acid on the mutagenicity of N-methyl-N-nitrosoguanidine and nitrofurans studied by SOS chromotest.

Mutagenicity inhibition of MNNG*, NFAA* as well as of nitrovin by ascorbic acid was observed. The influence of ascorbic acid on these compounds was parallelly studied also by spectroscopy. Mutagenicity inhibition was characterized by SOS chromotest. Inhibition degree was evaluated quantitatively introducing the coefficient of inhibition. The influence of ascorbic acid was expressed most markedly in MNNG. It has been found that the decrease of mutagenicity should be mainly caused by acceleration of MNNG decomposition by ascorbic acid.

Comparative chromosome examination of AKR mouse lymphoma after its i.p. transplantation with thymus- or spleen-derived lymphoma cells into hybrid mice.

A spontaneous AKR female mouse lymphoma was transplanted with its thymus cell suspension (10(6) cells i.p.) into AKR female mice. Lymph node cell suspension derived from one of the leukemic mice was injected (10(6) cells i.p.) into AKR females. The lymphoma cells "homing" to the thymus of one of the AKR females were suspended in Parker solution and 5 X 10(6) cells were given i.p. 5 (C3H X AKR) F3 hybrid mice (group 1). The lymphoma cells "homing" to the spleen of the same AKR female were also suspended, and 5 X 10(6) cells were injected i.p. into 5 (C3H X AKR) F3 hybrids (group 2). Chromosomes were prepared from the thymus and the spleen of two mice in both groups. The karyotypes derived from the hybrids of group 1 were compared to that of the group 2. It was found that the lymphoma cells "homing" to the thymus could be characterized by the trisomy of chromosome 15, while the lymphoma cells "homing" to the spleen had primarily the trisomy of chromosome 18. The results indicate that the thymus manifestation of the spontaneous AKR lymphoma is heterogeneous, and it contains at least two major subpopulations of the lymphoma cells.

Electrophoretic mobility patterns of immunologically phenotyped cells in different hemopoietic malignancies.

The electrophoretic mobility distribution along with the immunologic phenotype (rosette tests, surface membrane immunoglobulin determination and mainly the detection of differentiation and leukemia-associated antigens by monoclonal antibodies) have been studied in 120 patients with different hemopoietic cell malignancies and in a group of healthy donors. The aim of our study was to compare the electrophoretic mobility character of malignant cells with that of normal T and B lymphocytes and their immune phenotype. Normal peripheral blood lymphocytes showed the typical bimodal pattern of two clearly distinguishable populations of different electrophoretic mobilities, corresponding to T and B cells. In leukemia and lymphoma cells the sharp unimodal peak has appeared, which was attributed to monoclonal origin of these cells. Moreover, the electrophoretic mobility of cells in different hemopoietic cell malignancies reflected different cell lineages and maturation stages within the given lineage group. Utilizing of the cell electrophoretic mobility as an additional marker to the immunologic data for the characterization of leukemia and lymphoma cells is proposed.

The value of the short term predictive assay in the inductive treatment of patients with acute nonlymphoblastic leukemia.

Predictive capacity and clinical usefulness of the short term predictive assay (STPA) in the inductive treatment of patients with acute nonlymphoblastic leukemia (ANLL) was studied. Inductive treatment consisted of daunorubicin, arabinoside C and 6-thioguanine (TAD regimen). Leukemic cells of 20 previously untreated patients with ANLL were incubated in vitro with two doses of daunorubicin (1 microgram/ml and 10 micrograms/ml), arabinoside C (10 micrograms/ml and 100 micrograms/ml) and 6-thioguanine (10 micrograms/ml and 100 micrograms/ml). The 3H-thymidine as well as 3H-uridine uptake was measured in the treated and untreated cells. The highest predictive presence of the in vivo drug-sensitive disease was adequately reflected by the level of 3H-uridine incorporation suppression 30% of control value in the case of daunorubicin (concentration: 10 micrograms/ml) and 80% in the case of 6-thioguanine (concentration: 100 micrograms/ml). In the case of arabinoside C (concentration: 10 micrograms/ml) the limit of 3H-thymidine uptake depression was 20% of control value. It was rather difficult to define the indicative degree of precursors incorporation inhibition for prediction of the drug-resistant disease, because of low number of patients primary resistant to TAD regimen. No correlation was found between the degree of the pre-treatment DNA synthesis rates and the precursors uptake inhibition by the tested drugs.

Prognostic factors in Hodgkin's disease.

Between 1978-84 371 patients with HD were treated according to Protocol of LCG. 72.5% achieved complete, 10.8% partial remission and 16.7% were nonresponders. The evaluation of results was performed by PIL program from BMDP 81. The mean survival was 61 and disease-free survival (DFS) 60 months. The cumulative survival and DFS at 5 years were 65% and 68%, resp. Advanced HD, B symptoms, histopathology LD, male sex and age above 40 years influenced survival adversely. DFS decreased with CS III B, in males and age above 60 years significantly and in histopathology MC and LD with lack of significance. However, the size of mediastinal involvement was not a poor prognostic factor for survival, but influenced DFS. The prognostic value of major risk factors in HD was estimated by means of frequencies of their associations.

Antiimmunoglobulins of rheumatoid factor (RF) type in prediction of melanoma patients.

It was found out the titer of antiimmunoglobulins of the rheumatoid factor (RF) type in the sera of melanoma patients can be used in prediction of the development of the patients' status. High titers of antiimmunoglobulins of the RF type, determined by the latex-fixation test (LFT), indicate an unfavorable prognosis in patients with clinical manifestation of uncontrollable recurrence appearing from 1.5 to 2 years after occurrence of the LFT titer.

Lymphonodal response to sensitization with 2,4-dinitrochlorobenzene (DNCB) in laboratory rats.

The authors investigated the lymphonodal response to peripherally administered 2,4-dinitrochlorobenzene (DNCB). DNCB is known as a strong irritant binding to membrane proteins of the Langerhans cells of the skin. The conjugate is a potent neoantigen able to induce local tumor-destruction and raise cellular immune defence. White laboratory rats were used for the experiments. Their popliteal lymph nodes were investigated after sensitization with DNCB and i.c. injection of H3-thymidine and Patent Blue Violet at several intervals following the challenge with the hapten DNCB. Changes of weight of lymph nodes further uptake of H3-thymidine by the cells were investigated by autoradiography, while the content of H3-DNA indicating an increased DNA-metabolism due to blastic transformation of the small lymphocytes, was determined quantitatively by a liquid scintillation method. The data obtained speak for the usefulness of DNCB as an immunostimulant, comparable to BCG but of less toxicity.

Effect of hydroxyurea treatment on lysosomal membrane stability and enzyme latency in L5178Y cells in culture.

The stability of lysosomes prepared from hydroxyurea (HU) treated or untreated L5178Y cells and exposed to an HU-free iso- or hypoosmotic solution was compared. The data revealed that exposure of the cells to 1-10 mM HU for few hours tends to stabilize lysosomes. Partial protection against this effect could be afforded by catalase and superoxide dismutase. Under the same conditions of HU treatment lysosomal enzyme latency remained similar in HU treated and untreated cells.

Ovarian and endometrial endometrioid carcinoma: diagnostic implications of flow cytometry.

Surgical specimens from two patients with simultaneously occurring ovarian and endometrial endometrioid carcinoma were studied by flow cytometry (FCM) and routine histological examination. Tissue derived from the uterine and the ovarian lesion showed similar FCM curves in each case. The information obtained from FCM was more relevant than the study of frozen histological sections as far as confirming or ruling out a common histogenesis was concerned. Study of paraffin sections provided additional data, suggestive of metastatic spread from ovary to uterus in Case 1, and from uterus to ovary in Case 2.

In vitro monocyte maturation in patients with malignant melanoma and colorectal cancer--clinical significance.

In vitro monocyte maturation was studied in 52 malignant melanoma patients, 15 patients with colorectal cancer and 44 healthy donors. Index of maturation (IM) was in malignant melanoma patients 6.3 +/- 5.1%, in colorectal cancer 12.7 +/- 9.6%, and in healthy donors 40.4 +/- 18.0%. The difference between mean values in malignant melanoma patients and patients with colorectal cancer was significant when compared with healthy donors (p less than 0.001). The values in malignant melanoma patients decreased in accordance with stage of the disease and the difference between Stage I and Stage IV was significant (8.2 +/- 4.0% vs. 2.8 +/- 2.0%, p less than 0.01). In patients with colorectal cancer significant difference was established between operated patients and inoperable cases (21.4 +/- 10.0 vs. 5.1 +/- 1.9%, p less than 0.01). Monocyte maturation was influenced by the success of treatment. In patients with complete response the values were significantly higher than in patients with progression (11.7 +/- 6.7% vs. 2.2 +/- 1.5%, p less than 0.001). Autologous serum seems to inhibit the maturation process in vitro in cancer patients, while it apparently has no influence in healthy donors. When a response to treatment was achieved the previously low values of IM increased, while at progression a decrease was noted. The correlation between in vitro monocyte maturation and clinical factors leads to conclusion that the in vitro maturation reflects the in vivo process and may prove useful as a marker of tumor load or spread and may be a sensitive monitor of the treatment effect.

Zinc in human malignancies.

Serum zinc level (SZL) was determined in normal patients and in patients with primary tumors of different sites, with measurements made before and after therapy. Serum zinc concentration was depressed in some of the tumor types examined. In some cases serum zinc concentration increased again after successful treatment. Diagnostic and prognostic value of the serum zinc concentration in malignancy is discussed.

Procarbazin in advanced malignant melanoma.

Experience in applying procarbazin in the treatment of advanced malignant melanoma are very limited. Actually it has never been adequately tested as monotherapy. We administered procarbazin to 28 patients in a dosis of 100 mg/m2 on days 1 to 10 at 4-week intervals. As a result we obtained 25% remissions which suggest that procarbazin deserves greater attention in the above indication.

Gastric cancer in Poland--a decreased malignancy due to changing nutritional habits of the population.

The paper deals with the mortality pattern from stomach cancer over the period of last 20 years in Poland. The decrescent nature of gastric cancer was discussed in the light of data regarding the food consumption over several past decades and the results of the case-control study on stomach cancer and diet. In men the mortality rates dropped over 20 years by about 50% in younger and by about 40% in older age groups. Among women the drop in the rates was slightly faster than in men with the exception of age group 40-44 years. When examining the consumption of specific food products per capita over the period of last few decades, one has to note a marked increase in meat and drop in the consumption of cereals, as well as potatoes. The case-control study showed that the high risk of stomach cancer run the people with low level of vegetable and fruit consumption. Analysis performed confirmed the downward trend in stomach cancer rates in Poland over the last 20 years and explains the pattern observed by the marked changes in the nutritional habits of the population at large.

Studies on viral etiology of angioimmunoblastic lymphadenopathy.

Plasma samples of patients with angioimmunoblastic lymphadenopathy (AIBL) were tested for anti-HTLV antibodies and for interferon content. Out of 12 patients 4 had antibodies to HTL-III. Two of these plasma samples contained antibodies reacting with HTLV-I and HTLV-II, too. Activated interferon (IFN) system was found in patients with clinical remission of AIBL, as it was detected by IFN titration in their plasma samples. Data suggest the etiological role in AIBL of virus(es) related to the HTLV family.

Efficacy of combination chemotherapy including Platidiam in the treatment of malignant testicular tumors.

Thirty-seven patients were treated for advanced malignant testicular tumors with combined chemotherapy including cis-platinum (Platidiam). A complete regression was achieved in 23 patients (62%). No severe side effects were observed. Also in this study the effectivity of cis-platinum in the treatment of testicular tumors was proved.

The effect of nondamaging intensity laser irradiation on the immune system.

Data on in vitro studies of the effect of nondamaging laser irradiation on peripheral blood lymphoid cells of donors and cancer patients are reported including the results of experimental investigations carried out in patients with breast cancer and malignant melanoma. Laser irradiation is demonstrated to potentiate the immunological parameters both in healthy individuals and patients, the effect being more marked in cancer patients. A different action of red and blue laser irradiation on the immune system was revealed. The evidence obtained may be useful in working out the immunotherapeutic methods in cancer management.

A simple and rapid method for the isolation of human alpha-fetoprotein from human cord serum.

A description is given of a simple and time-saving method for the isolation of human alpha-fetoprotein (AFP) by means of liquid chromatography. The isolation was made in three steps: Step 1--affinity chromatography on column CNBr-Sepharose 4 B coupled with sheep antibody against human alpha-fetoprotein. AFP was eluted from the column by a change pH. Step 2--application of AFP to column Blue-Sepharose CL-6B for the removal of albumin remnants. Step 3--additional purification of the AFP preparation on column Con-A-Sepharose. AFP was eluted from the column by using methyl-alpha-D-glucepyranoside gradient. The isolation yield was 20%. The purity of the AFP preparation was satisfactory for RIA.

Low-molecular weight inhibitory factor released in vitro by murine L5178Y leukemic cells.

A factor released spontaneously by murine leukemic lymphoblasts L5178Y into an isotonic nonnutrient saline during short-term incubation at 37 degrees C is described. The factor is dialysable and its molecular weight was found to be below 500. The factor exhibits very high thermal stability and is not inactivated by UV irradiation. In L5178Y cells it strongly inhibits biosynthesis of RNA and proteins by interfering with precursors uptake as well as with their incorporation into respective macromolecules. Its action is reversible, abolished by cell washing. In spite of its inhibitory effect on biosynthesis of RNA and proteins it does not inhibit cell growth in vitro.

Significance of CEA, AFP, SP1 and HCG as tumor markers in ovarian carcinoma.

The significance of CEA, AFP, HCG and SP1 determinations in 85 patients with ovarian cancer were analyzed from the point of view of evaluation of the clinical status of the patients. On the basis of the trials it was stated that a satisfactory correlation between the extension of the tumor and the serum CEA level was observed in 26.6% of the cases. The efficiency of monitoring by systematic CEA tests was found to be 17.3%. The determination of AFP and HCG is useful only in cases of tumors with specific histological structures and in problems of differential diagnostics. The testing of SP1 is of no value in clinical practice.

High degree of resistance to 5-aza-2'-deoxycytidine in L1210 cells in vitro associated with almost complete loss of deoxycytidine kinase activity.

In L1210 mouse leukemic cells the resistance towards 5-aza-2'-deoxycytidine was developed in vitro by stepwise selection in increasing concentrations of this drug. According to the respective IC50 values the variant cell line was at least 7000-fold more resistant to 5-aza-2'-deoxycytidine, 3300-fold to arabinosyl 5-azacytosine and 20,000-fold to arabinosyl cytosine in comparison to the parental cell line. The uptake of radioactivity derived from 5-aza-2'-deoxycytidine in variant cells was depressed by 95% as compared to the parental strain and no uptake occurred in case of 2'-deoxycytidine. The phosphorylation of this natural precursor and of its analog in the presence of the cell-free system derived from the variant cell line was nearly completely abolished. In resistant cells the amount of DNA 5-methyl-cytosine relative to cytosine was decreased by 24-44%.

Non-Hodgkin's lymphoma in children. Results of therapy.

Twenty-one children with non-Hodgkin's lymphoma (NHL) entered intensive chemotherapy study according to the protocol 6481. Sixteen patients with extraabdominal involvement achieved long-lasting remission, 15 of them have been disease-free for 17 till 77 months. Five children suffered from intraabdominal NHL, 4 of them expired because of progression of the disease within 4 months after diagnosis, 1 patient has been disease-free for 43 months. Actuarial disease-free survival is 0.76 for the whole group, 0.8 for those with extraabdominal involvement and 0.20 for children with intraabdominal forms of non-Hodgkin's lymphoma. Protocol 6481 is highly effective for extraabdominal forms of childhood non-Hodgkin's lymphomas but insufficient for abdominal forms.

Product of the src gene is not phosphorylated in avian sarcoma virus B77 infected but untransformed human cells.

Human embryo cells infected with the avian sarcoma virus B77 [Hu(B77)] but untransformed, contain the whole rescuable virus genome integrated in the host cell DNA. The cellular DNA induced the transformation of the infectious B77 virus after transfection of chicken cells. In the Hu(B77) cells the src gene product was expressed as a 58 kD protein which possessed phosphokinase activity but was not phosphorylated in comparison with the src production of B77 transformed rat cells RBI in which p60src is expressed. The addition of vanadate to tissue culture fluid reversibly elicited cell transformation in both control and virus infected human cells, but did not influence phosphorylation of the src gene product. The secretion of phosphoproteins in the investigated cells was different. Whereas transformed rat cells released a 62 kD transformation related phosphoprotein, the human cells did not. In tissue culture fluid from Hu(B77) cells an elevated amount of a 22 kD phosphoprotein was found in comparison with control cells. The implications of these findings are discussed with respect to the role of the v-src gene product in malignant cell transformation.

Delayed hypersensitivity to polyoma and "self" antigens in syngeneic mice.

Using the tail swelling test, delayed-type hypersensitivity (DTH) to tumor and self antigens accompanying the development of syngeneic polyoma tumor in CBA mice was observed. The soluble polyoma cell-surface antigen contained both polyoma and H-2k specificities and induced weak proliferation response of spleen cells in the presence of Il-2 in vitro. Appearance of a measurable tail swelling reaction at the side of the midtail, subcutaneous injection of the eliciting antigen not earlier than at 16th hour, maximum of swelling after 24 hours and the results of histological examination proved that the swelling was caused by a typical DTH to the antigens. Maximum DTH to both antigens occurred in the mice 6 days after polyoma cell transplantation (about 3 days before the appearance of palpable tumor) and weakened as the tumor progressed. DTH activity was transferred by spleen T lymphocytes to naive recipients. Five-day restimulation of splenocytes from polyoma transplanted donors with antigen in the presence of Il-2 led to an increase of antigen-lymphocytes affinity but resulted in a decrease of DTH activity of these cells. The mechanisms of these processes are still discussed.

Bovine leukemia virus: isolation and characterization of nonproducer cell clones.

Several single-cell clones were isolated from a fetal lamb kidney cell line (FLK) persistently infected with bovine leukemia virus (BLV). The majority of isolated cell clones were virus productive, several were nonproductive based on the determination of the activity of reverse transcriptase and production of 3H-uridine labeled virus particles. Two nonproductive clones, NP-1 and NP-2, were further characterized in comparison with the virus productive cells. All clones contained three integrated BLV proviruses in the nonproducer cells. The virus specific mRNAs were expressed both in the virus productive and nonproductive cells. The virus-specific protein products were found different in the nonproducing cells. The gag pol precursor Pr145 was missing in NP-1 cells, in NP-2 its Mr was 120 only. In NP-2 cells the precursors Pr70gag and Pr45gag were absent. The env precursor gPr72 and both of the two glycoproteins were detected. The nonproductive cells NP-2 produced mainly env gene products, therefore they were tested as a potential material for anti-BLV vaccine. The NP-2 cells after inoculation to rats and cattle were able to induce formation of neutralizing antibodies directed against the env gene products.

Provirus integration of bovine leukemia virus into DNA of infected human myeloma cells.

Sixty clones of bovine leukemia virus-infected B-human myeloma ARH77 cells were isolated. The DNAs of all clones were examined for BLV provirus integration by Southern blotting analysis. Proviral sequences were found in DNAs of two clones. One of them (clone I B3) contained one proviral copy with a deletion of approximately 5.5 kb; the other one (clone I F9) carried three integrated proviruses. Viral proteins of 70,000, 42,000 and 35,000 M. W. were found in extracts of clone I F9. In clone I B3 only the 70,000 M.W. protein was detected.

The excretion of urinary enzymes, proteins and creatinine in patients receiving cisplatinum.

The sensitive parameters of tubular nephrotoxicity during 4-day chemotherapy with new combination schedule of cisplatinum and 5-fluorouracil were followed. The determinations of tubular enzymes beta-glucuronidase (GRS) and gamma-glutamyltransferase (GMT) in 24 hours urine, the excretion of creatinine and proteinuria were assayed before therapy and during 4 consecutive days of treatment. We recorded the significant increase of protein excretion and only slight increase of GMT and GRS activities after chemotherapy. The decrease of the creatinine excretion on the 3rd day of therapy was not statistically significant. Simultaneously followed serum creatinine and urea levels proved the elevation of creatinine only in 1 patient (the 1st grade according WHO classification). Our results suggest that the performed regime of chemotherapy produces only very low nephrotoxicity comparing to the 1-day administration of cisplatinum. These conclusions are confirmed also by the former examinations of GRS activities in cisplatinum therapy.