6 research outputs found
Multicenter fresh frozen tissue sampling in colorectal cancer: does the quality meet the standards for state of the art biomarker research?
The growing interest in the molecular subclassification of colorectal cancers is increasingly facilitated by large multicenter biobanking initiatives. The quality of tissue sampling is pivotal for successful translational research. This study shows the quality of fresh frozen tissue sampling within a multicenter cohort study for colorectal cancer (CRC) patients. Each of the seven participating hospitals randomly contributed ten tissue samples, which were collected following Standard Operating Procedures (SOP) using established techniques. To indicate if the amount of intact RNA is sufficient for molecular discovery research and prove SOP compliance, the RNA integrity number (RIN) was determined. Samples with a RIN < 6 were measured a second time and when consistently low a third time. The highest RIN was used for further analysis. 91% of the tissue samples had a RIN ā„ 6 (91%). The remaining six samples had a RIN between 5 and 6 (4.5%) or lower than 5 (4.5%). The median overall RIN was 7.3 (range 2.9ā9.0). The median RIN of samples in the university hospital homing the biobank was 7.7 and the median RIN for the teaching hospitals was 7.3, ranging from 6.5 to 7.8. No differences were found in the outcome of different hospitals (p = 0.39). This study shows that the collection of high quality fresh frozen samples of colorectal cancers is feasible in a multicenter design with complete SOP adherence. Thus, usin
Determination of Ki-67 defined growth fraction by monoclonal antibody MIB- I in formalin-fixed, paraffin-embedded prostatic cancer tissues
The applicability of MIBā1, a monoclonal antibody directed against the Kiā67 antigen, was studied in the PCā82 and LNCaP prostatic tumor models at various levels of proliferative activity. Statistically significant correlations were found in LNCaP cultures between Kiā67 and MIBā1 scores (r = 0.84, P < 0.001), and in PCā82 tumors between MIBā1 scores and paraffin tissue Kiā67 (pKiā67) (r = 0.90, P < 0.001), frozen tissue Kiā67 (fKiā67) (r = 0.86, P < 0.001), and BrdU uptake (r = 0.70, P < 0.001), respectively. pKiā67 scores were double the fKiā67 scores, which may be due to methodological differences. MIBā1 scores exceeded both the fKiā67 and pKiā67 scores. The affinity of MIBā1 for the antigen is much higher than the affinity of Kiā67, which may explain the differences. MIBā1 is a promising means of evaluating the presence of only minute amounts of the Kiā67 antigen in paraffināembedded human tumor material, especially in relatively slowly growing tumors
Kinetics of neuroendocrine differentiation in an androgen-dependent human prostate xenograft model
It was previously shown in the PC-295 xenograft that the number of
chromogranin A (CgA)-positive neuroendocrine (NE) cells increased after
androgen withdrawal. NE cells did not proliferate and differentiated from
G0-phase-arrested cells. Here we further characterized NE differentiation,
androgen receptor status, and apoptosis-associated Bcl-2 expression in the
PC-295 model after androgen withdrawal to assess the origin of NE cells.
PC-295 tumor volumes decreased by 50% in 4 days. Intraperitoneal
bromodeoxyuridine (BrdU) incorporation and MIB-1 labeling decreased to 0%,
and the apoptosis was maximal at day 4. Androgen receptor expression and
prostate-specific antigen (PSA) serum levels decreased rapidly within 2
days. The number of NE cells increased 6-fold at day 4 and 30-fold at day
7. Five and ten percent of the CgA-positive cells were BrdU positive after
continuous BrdU labeling for 2 and 4 days, respectively. However, no MIB-1
expression was observed in CgA-positive cells. NE cells expressed the
regulated secretory pathway marker secretogranin III but were negative for
androgen receptor and Bcl-2. Bcl-2 expression did increase in the non-NE
tumor cells. In conclusion, androgen withdrawal leads to a rapid PC-295
tumor regression and a proliferation-independent induction of NE
differentiation. The strictly androgen-independent NE cells that were
still present after 21 days differentiated mainly from G0-phase-arrested
cells
Expression and prognostic value of Wilms' tumor 1 and early growth response 1 proteins in nephroblastoma
Wilms' tumor is one of the most common solid tumors of children. The
protein product of the tumor-suppressor gene, Wilms' tumor 1 (WT-1), binds
to the same DNA sequences as the protein product of the early growth
response 1 (EGR-1) gene. There is experimental evidence that EGR-1 is
involved in controlling cell growth. The expression of both genes in
Wilms' tumor was studied by others, mainly at the mRNA level. The present
study evaluates the prognostic value of WT-1 and EGR-1 in 61 Wilms' tumors
of chemotherapeutically treated patients at the protein level, using an
immunohistochemical approach. WT-1 was expressed in normal kidney tissues
and in the blastemal and epithelial component of Wilms' tumor, whereas
stromal tissue was negative. EGR-1 was expressed in normal kidney tissues
and in the three main cell types of Wilms' tumor. In 59 and 56% of Wilms'
tumor, the blastemal cells stained for WT-1 and EGR-1, respectively. The
blastemal expression of WT-1 and EGR-1 and the epithelial expression of
WT-1 were statistically significantly correlated with clinical stage. WT-1
immunoreactivity correlated with EGR-1 expression. Univariate analysis
showe
Cryo-Gel embedding compound for renal biopsy biobanking
Optimal preservation and biobanking of renal tissue is vital for good diagnostics and subsequent research. Optimal cutting temperature (OCT) compound is a commonly used embedding medium for freezing tissue samples. However, due to interfering polymers in OCT, analysis as mass spectrometry (MS) is difficult. We investigated if the replacement of OCT with Cryo-Gel as embedding compound for rena
Androgen deprivation of the PC-310 [correction of prohormone convertase-310] human prostate cancer model system induces neuroendocrine differentiation
Neuroendocrine (NE) cells are androgen-independent cells and secrete
growth-modulating neuropeptides via a regulated secretory pathway (RSP).
We studied NE differentiation after androgen withdrawal in the
androgen-dependent prostate cancer xenograft PC-310. Expression patterns
of chromogranin A, secretogranin III, and prohormone convertase-1 were
analyzed at both protein and mRNA level to mark the kinetics of NE
differentiation both in vivo and in vitro. PC-310 tumor-bearing nude mice
were killed at 0, 2, 5, 7, 14, and 21 days postcastration. PC-310C
cultures initiated from collagenase-treated tumor tissue could be
maintained up to four passages, and androgen-deprivation experiments were
performed similarly. PC-310 tumor volumes decreased by 50% in 10 days
postcastration. Proliferative activity and prostate-specific antigen (PSA)
serum levels decreased to zero postcastration, whereas PSA levels in
PC-310C culture media first decreased and subsequently increased after 5
days. In vivo, androgen receptor (AR) expression decreased initially but
returned to control level from 5 days postcastration on. CgA,
secretogranin III, and secretogranin V expression increased in vivo from 5
days postcastration on. Subsequently, prohormone convertase-1 and peptidyl
alpha-amidating monooxygenase as well as the vascular endothelial growth
factor were expressed from 7 days postcastration on, and, finally, growth
factors such as gastrin-releasing peptide and serotonin were expressed in
a small part of the NE cells 21 days postcastration. The PC-310 tumors did
not show colocalization of the AR on the NE cells in the tumor residues
after 21 days. As in the PC-310 xenograft, NE differentiation was induced
and AR expression relapsed after prolonged androgen suppression in
PC-310C. For PC-310C cells, this relapse was associated with the secretion
of PSA. PC-310C is the first culture of human prostatic cancer cells
having the NE phenotype. The PC-310 model system is a potential
androgen-dependent model for studying the role of NE cells in the
progression of clinical prostate cancer. Androgen deprivation of
NE-differentiated prostate cancer may induce the formation of both NE- and
AR-positive dormant tumor residues, capable of actively producing NE
growth factors via a RSP, possibly leading to hormone refractory disease