Real-world analysis of teclistamab in 123 RRMM patients from Germany.

Teclistamab, a B-cell maturation antigen (BCMA) × CD3 directed bispecific antibody, has shown high response rates and durable remissions in the MAJESTEC-1 trial in patients with relapsed and refractory multiple myeloma (RRMM). We retrospectively assessed efficacy and tolerability in 123 patients treated at 18 different German centers to determine whether outcome is comparable in the real-world setting. Most patients had triple-class (93%) or penta-drug (60%) refractory disease, 37% of patients had received BCMA-directed pretreatment including idecabtagene vicleucel (ide-cel) CAR-T cell therapy (21/123, 17.1%). With a follow-up of 5.5 months, we observed an overall response rate (ORR) of 59.3% and a median progression-free survival (PFS) of 8.7 months. In subgroup analyses, we found significantly lower ORR and median PFS in patients with extramedullary disease (37%/2.1 months), and/or an ISS of 3 (37%/1.3 months), and ide-cel pretreated patients (33%/1.8 months). Nonetheless, the duration of response in ide-cel pretreated patients was comparable to that of anti-BCMA naive patients. Infections and grade ≥3 cytopenias were the most frequent adverse events. In summary, we found that teclistamab exhibited a comparable efficacy and safety profile in the real-world setting as in the pivotal trial.

Reactive astrogliosis after basic fibroblast growth factor (bFGF) injection in injured neonatal rat brain.

Reactive gliosis was revealed by immunocytochemistry using antibodies against the glial fibrillary acidic protein (GFAP) after a stab or an electrolytic lesion administered to the cerebral cortex, corpus callosum, striatum, or hippocampus of a 6-day-old rat. The intensity of the gliosis was about the same in the various structures injured and did not change with the delay of 3, 7, or 20 days between the injury and the sacrifice of the animals. When basic fibroblast growth factor (bFGF) was injected in the lesion locus just after the lesion was performed, it resulted (as soon as 3 days after injury) in a strong astrogliosis that was enhanced after a delay of 7 days, the astrocytes in the lesion area exhibiting enlarged cell processes and intense GFAP-positive immunoreactivity. After a delay of 20 days, the astrocytes were not dispersed any more but packed in three or four layers along the borders of the lesion, thus reducing its extension. This suggests a possible role for bFGF in promoting scar formation following brain injury.

Embryonic spontaneous motility after acute and chronic administration of kainate.

Changes in spontaneous motility after the acute and chronic administration of kainate were studied in 11- to 19-day-old chick embryos. 1. After acute administration, kainate (20 mg/kg e.w.) already depressed motility in 11-day-old embryos. From the 17th day it induced explosive activation of embryonic motility, but never in chronic spinal preparations. 2. The chronic administration of kainate (2.56 +/- 0.62 mg/kg e.w./24 h) reduced embryonic motor activity. The effect already developed after administering kainate from the 4th to the 8th day of incubation. Prolonged administration made no important difference to the results. Chronic administration was followed by histopathological changes in the nervous tissue. These were mainly of an oedematous type and affected the glia and the brain capillaries, whereas pyknotic changes were found in the large neurones. 3. The results showed that the CNS is already sensitive to the neurotoxic effect of kainate from the early stages of embryogenesis and that the picture of the reaction of the embryonic CNS is closely correlated to the degree of maturation.

Correlation between changed biophysical properties of surface membranes and embryonic brain cell adhesivity. Influence of detergents, fixation, EGTA, colchicine, vinblastine, increased K+, ouabain and DMA on the primary cellular adhesivity of embryonic brain cell.

Embryonic mouse brain cells were rotated for 120 min and cellular adhesivity was tested under normal conditions and in the presence of substances which change the membrane properties. A marked decrease of cellular adhesivity (but not complete inhibition) was recorded in the presence of anionic detergents, while fixation of cells caused only non-significant inhibition Colchicine (1 mumol/l) and vinblastine (10 micrograms/ml) did not significantly affect the adhesivity. Increased external K+ (10 mumol/l) and ouabain (10 mmol/l) were also without a significant effect, however, EGTA (0.1 and 0.01 mmol/l) inhibited the adhesivity significantly. 2,3 dimethyl maleinic anhydride (DMA) which removes a part of the positive charge, caused a slight decrease of adhesivity. It is suggested that the primary adhesivity of brain cells is dependent upon the structural integrity of surface membranes, while the organization of the tubular system does not play a significant role. Isotonic concentration of monovalent cations is optimal for adhesivity and an increased concentration of external K+ or ouabain did not affect adhesivity significantly.

Influence of increased concentration of Ca2+, La3+, PVP and urea on primary adhesivity of embryonic brain cells. Ultrastructural pattern of adhering cells.

With the help of a previously described experimental arrangement the influence of increased external concentration of Ca2+, La3+, PVP and urea was tested on the initial stages of brain cell adhesivity and its kinetics. Urea, an inhibitor of hydrogen bonds, significantly inhibited the adhesivity of the treated cells. PVP significantly increased cell adhesivity. The adhesivity was enhanced and speeded up by increased concentrations of Ca2+ and La3+. It is evident that the membrane surface potential, zeta potential and formation of H+ bonds and bridges are highly important for cellular adhesivity. EM control of freshly dissociated cells disclosed that a part of the cells had been damaged. According to the ultrastructural organisation, the surface membrane is damaged to a small or greater extent. Intercellular contacts were formed in vitro either between non damaged surfaces of membranes, or between fragments of membranes, or contacts were mediated by membrane debris. Because cellular debris disappeared during rotation from single adhesive complexes, it is probable, that disrupted membranes are used for restoration of membranes, or serve as a metabolic substrates, or are catabolized.

The organization and differentiation of nervous cells in tissue cultures (a minireview).

Single stages of histiotypic formation from dissociated embryonic brain cells are described. The first stage, i.e. primary adhesion, is a phenomenon depending mainly on physicochemical features of the adhesive system. The composition of the membrane glycoprotein coat was studied during membrane regeneration and formation of cellular contacts. At the beginning single terminal sugar components and negative sialic acid residues are distributed non-homogeneously over the membrane according to the ligation of lectins. During the formation of cellular contacts this non-homogeneity progressively disappears, the thickness of the layer is reduced, however, the density of single markers increases. The highest increase of both density and layer thickness during the phase of membranes reparation occurs when the negative surface groups are labelled with cationized ferritin. The sorting out process was studied in mixed cultures as a stage of final specific recognition. It is assumed that the reaggregation of cells is a multistep process depending on maturation of the glycoprotein coat, characterized by multiple cellular adhesion and deadhesion, completed by specific recognition and fixation of recognized cells.

The effect of short-term temperature changes on the adhesivity of nerve cells. Participation of DNA molecules on non-specific cellular adhesion.

Brain cells from 16 to 18-day-old mice embryos were dissociated by mild trypsinization and sieving. Immediately after dissociation the cells were preincubated in a PBS solution at -6 to +54 degrees C for 3 and 20 min. After this preincubation cells were rotated for 60 min at 37 degrees C in the PBS solution. Cellular adhesivity was estimated during this time period and EM pictures of organized in vitro aggregates after 24-28 h were taken. In a separate series of experiments, freshly dissociated were treated with DNAase before the rotation procedure. Preincubation in a cold or a warm medium did not alter the inhibition of cellular adhesivity significantly. Distinct inhibition of cellular adhesion was observed in cells preincubated above 53 degrees C. Adhesion was also inhibited below -5 degrees C, however, this effect was mainly dependent on the rate of freezing and thawing. Digestion of dissociated cells with DNAase (20 micrograms/ml) decreased cell adhesion. At 37 degrees C the adhesivity decreased by about 20%. Aggregates of cells preincubated at 0 degrees C for 20 min did not exhibit marked EM changes after 24-28 h in vitro. The present results have shown the rather high resistance of molecules responsible for cellular adhesion and its reversibility to temperature changes. Furthermore, non-specific cellular adhesion was shown on physically active DNA molecules.

Morphological maturation and survival of chicken and rat embryonic neurons on different culture substrata.

Neuronal cells from chicken and rat embryonic cerebral hemispheres were plated at a low cell concentration and cultured either on collagen or on a supporting continuous glial layer for periods of up to 21 days. The glial layer was either homologous or heterologous with regard to the animal species; the survival and maturation of the neuronal cells in these different conditions were investigated by light and electron microscopy. Neuronal cells cultured on collagen formed aggregates similar to those formed by neuronal cells plated at high cell density as described in a previous paper; a few aggregated neurons formed processes after 24 h and, only after 48 h of culture, more fibres had developed; the majority of the cells progressively degenerated between days 7 and 21 of culture. In contrast to this, neuronal cells cultured on a supporting glial layer, whether homologous or heterologous, progressively differentiated: neuronal perikarya remained well separated from each other and many processes were already formed after 24 h; later on, networks of fibres developed. At the electron microscopic level, microtubules and neurofilaments were present at a high density in the cells and fibres; immature synapses could be found, but infrequently. Differentiated cells were represented mostly by neurons; oligodendroglial cells were absent, and myelinated fibres could not be detected. The highest positive effect on the maturation and survival of neuronal cells was observed in the presence of a layer of glial cells from the same species. These results emphasize the essential role of glial cells for the neuronal maturation in the absence of contact between neuroblasts.

Inhibitory effect of tunicamycin on the adhesivity and aggregation of embryonic brain cells.

Tunicamycin (TM) - (0.1, 1.0 and 10 micrograms/ml) inhibits insignificantly the adhesivity of embryonic mice brain cells during the first 120 min of incubation. The effect is not dose dependent. The concentration of 10 micrograms/ml added at the onset of experiments has a drastic effect during the time period in which cell regeneration, cell movement and formation of aggregates occurs. Up to the 5th day in vitro (DIV), aggregation is completely inhibited and disrupted parts of cells are mostly present in the medium. The concentration of 1 microgram/ml is less effective, and 0.1 microgram/ml is practically without effect. EM analysis shows that tunicamycin (10 micrograms and 1 microgram/ml) diminish the regeneration and integrity of plasmatic membranes, 10 micrograms/ml of tunicamycin destroys cytoplasmatic organelles which is probably the cause of the decline of cellular regeneration and of aggregate formation. Tunicamycin (10 micrograms/ml), if added to the already formed aggregates evokes their disintegration and lower doses (1 microgram/ml) liberated cells from aggregates into the medium.

Ultrastructural and protein synthetic changes in embryonic brain: cells aggregated at 0 degree C.

Cells dissociated from brains of 16 to 18 day-old mice embrya were rotated at 37 degrees C and 0 degree C for 7--8 days. While cells aggregated at 37 degrees C formed compact aggregates, cells aggregated at 0 degree C were found in clusters or were randomly distributed. Cells aggregated in the cold did not differ markedly from the controls in their ultrastructural organisation till the 2--3 day in vitro (DIV). Later, significant structural changes, such as distention of cytoplasmic membranes, destruction of mitochondrial membranes, disappearance of ribosomes, shrinkage of nuclei and disturbance of cytoplasmic membranes were apparent. On the 6--7 DIV, groups of cells were separated by a distance of 100 nm and more, and large parts of their cytoplasm disappeared and outside cell perikarya fragments of membranes appeared forming dense debris. However, even at this period some cells were found which did not show signs of degeneration. Protein synthetic activity in aggregated cells increased linearly at 37 degrees C till 7 DIV, whereas in cells aggregated at 0 degree C an inhibition of about 34% was found at 4 DIV and at 7 DIV the curve of 14C leucine incorporation declined almost to zero. It is thus evident that cells aggregated at 0 degree C maintain an almost normal ultrastructural pattern during the first days of cultivation and only protein synthetic activity is lowered. Cellular membranes, damaged during the dissociation partly regenerated even at 0 degrees C and membraneous contacts were formed between several cells.

Enzyme activity pattern in developing mouse brain in situ in embryonic brain aggregated cells at 37 degrees C and 0 degree C.

In aggregates of nervous tissue, cultivated for 1--7 days at 0 degree C and 37 degrees C, respectively, the activities of seven enzymes of energy liberating metabolism were estimated, in order to evaluate their metabolic "profiles" and changes during cultivation. The enzymes used as markers of different pathways of energy liberation from substrates were: lactate dehydrogenase - LDH - (EC 1.1.1.27), triose-3-phosphate dehydrogenase - TPDH - (EC 1.2.1.12), glycerol-3-phosphate dehydrogenase - GPDH - (EC 1.1.1.8), hexokinase - HK - (EC 2.7.1.1.), malate:NAD dehydrogenase - MDH - (EC 1.1.1.37), citrate synthase - CS - (EC 4.1.3.7), and 3-hydroxyacetyl CoA dehydrogenase - HOADH - (EC 1.1.1.35). During the cultivation, some changes in the metabolic "profiles" were observed. Although some of these changes as well as the differences between the cultivation at 0 degree C and 37 degrees C, were statistically significant, they were not greater than the variations between different samples of any tissue taken at different times. They were not, therefore considered to be of major significance. However, all the aggregates exhibited "profiles" characteristic for the nervous tissue, with relatively very high activity of HK, high activity of MDH and CS (carbohydrate breakdown) and low activity of GPDH and HOADH (lipid catabolism).

Reaggregation of human, chick, and human embryonic brain cells. Factors influencing the formation of a histiotypic unit.

1. Aggregation of embryo human, mouse, and chick brain cells was studied. The optimum age interval of donors from different species was determined. 2. The significance of different dissociation procedures (mild trypsinisation followed by sieving, trypsinisation + DNA digestion, mechanical dissociation in 1 or 2 steps, and Ca2+ chelation by EGTA) for the rate of aggregation was estimated. A significant reduction of aggregation was observed after one step mechanical dissociation. Nonspecific adhesion of cells on DNA molecules was found only during the first stages of aggregation. 3. The curve of aggregation kinetics follows the curve of floculation kinetics. 90% free cells disappear from the medium after 2 h of aggregation and a large number of microaggregates are formed which condense after 20 to 24 h into compact aggregates. The time course of aggregation was similar for all cells dissociated by different means. Small differences in the rate of aggregation, caused by dissociation procedures, were apparent only during the first stages of aggregation. 4. The histiotypic unit formed by aggregation of human, mouse, and chick embryo brain cells exhibits some common and some specific features. During aggregation a multiple structural reconstruction takes place and a limited number of cells are exchanged or sorted out from aggregates into the medium. 5. The structural organisation of aggregates from differently dissociated cells differs in several aspects. This indicates that membrane surface structures are influenced differently by dissociation and behave differently during distinct stages of aggregation.

The effect of dibutyryl cyclic AMP on cultivated glial cells from corpus callosum of 30-day-old rats.

1) Fragments of corpus callosum of 30-day-old rats were cultivated in Rose chambers. Mature astrocytes and predominantly differentiated oligodendroglial cells are present in the corpus callosum of rats at this age. 2) In vitro, astrocytes dedifferentiated into epithelial--like flattened cells, which highly adhered to the surface. Cells exhibited signs of phagocytosis, and their ultrastructure differed from differentiated cells, especially by the absence of microtubules and microfilaments. 3) After dibutyryl cyclic AMP (db-c-AMP) application, epithelial--like cells became rounded and multipolar. Cells were less liable to adhere to the surface. Bundles of microfilaments and microtubules appeared in cell perikarya and their processes. 3--4 hour after application, the morphological changes were well developed and disappeared after 24--48 hours. 4) It is being suggested that the increased intracellular concentration of db-c-AMP induced the formation of microfilamentous structures. The formation of the cyto-skeleton has a direct influence on the shape of cells and appearance of processes and cell adhesivity. 5) These results have demonstrated the high structural plasticity of mature astrocytes, their dedifferentiation in vitro and convertion into mature cells after db-c-AMP application.

Structure and ultrastructure of cultivated glial cells from Corpus callosum.

1. Explants and dissociated cells from Corpus callosum (c. c.) of rats and rabbits were cultivated in Petri dishes and Rose chambers. 2. Different types of glial cells were found in the cultivated Corpus callosum (c. c.) explanted from 12 days old rats: a) adendritic glial cells, typical for migrating oligodendroglial cells, b)-migrating large, nondifferentiated astrocytes with pronounced phagocytosing activity, c) macro- and microglial cells which differentiated during cultivation. 3. The population of differentiated glial cells is mostly composed of oligodendroglia, less of astrocytes and microglial cells are rare. 4. Differentiation of dissociated cells from c. c. in homogenous and mixed population was studied. The appearance of first processes of macroglial cells is postponed to 6 to 10 days of cultivation. No substantial difference was observed between homogenous and mixed population. A higher incidence of macrophages was observed in the later. 5. Glial cells differentiate surrounded by degenerated nerve fibers and myelin, exhibiting phagocytoses and cleaning reaction.

Differentiation of corpus callosum glial cells and factors influencing their maturation in vitro.

1. Explants of Corpus callosum (c. c.) from 12-day-old rats were cultivated under different experimental conditions. 2. Migration and differentiation is activated by the presence of neighbouring explants, toward which glial cells predominantly migrate. Glial cells migrate if closely adhering to the supporting collagen and the process of differentiation is enhanced by presence of underlying cell layers. 3. Migratory activity of glial cells decreases and is delayed with age of donors. Migrating cells have a similar appearance as in cultures from 12 days old donors. The presence of immature types of glial cells in c. c. of adult animals was proved. 4. Glucose was found to be an adequate metabolical substrate, utilisation of glucose being lower than in cultivated neurons. In the absence of glucose or serum in the medium, neither migration nor differentiation of glial cells was observed. 5. The addition of embryonal extract and embryonic brain extract enhanced only initial stages of cell migration and differentiation.