Content deleted Content added
Adding {{linkless}} template to orphan article |
m mu not micro per MOS:NUM#Specific units and Unicode compatibility characters (via WP:JWB) |
||
| (173 intermediate revisions by 69 users not shown) | |||
Line 1:
{{More citations needed|date=December 2007}}
[[File:LNCaP.jpg|thumb|Representative phase-contrast image of LNCaP cells. Scale bars represent 100 μm.]]
[[File:Comparison of cancer cell lines.png|thumb|Properties of common PCa cell lines]]
'''LNCaP''' cells are a [[cell line]] of human cells commonly used in the field of [[oncology]]. LNCaP cells are [[androgen]]-sensitive human [[prostate]] [[adenocarcinoma]] cells derived from the left supraclavicular lymph node [[metastasis]] from a 50-year-old caucasian male in 1977. They are adherent epithelial cells growing in aggregates and as single cells.<ref name="pmid6831420">{{cite journal |vauthors=Horoszewicz JS, Leong SS, Kawinski E, Karr JP, Rosenthal H, Chu TM, Mirand EA, Murphy GP | title = LNCaP model of human prostatic carcinoma | journal = Cancer Res | volume = 43 | issue = 4 | pages = 1809–18 |date=April 1983 | pmid = 6831420 }}</ref>
One major obstacle to conducting the most clinically relevant [[prostate cancer]] (PCa) research has been the lack of cell lines that closely mimic human disease progression.<ref name="pmid8168083" /> Two hallmarks of metastatic human prostate cancer include the shift of aggressive PCa from androgen-sensitivity to an Androgen Insensitive (AI) state, and the propensity of PCa to metastasize to bone.<ref>{{cite web|last1=Fanelli|first1=Alex|title=LNCaP Cell Line: human prostate adenocarcinoma cell line|date=2016|url=http://www.lncap.com/|access-date=3 December 2017}}</ref> Although the generation of AI cell lines has been quite successful as demonstrated in the “classic” cell lines [[DU145]] and [[PC3]], the behavior of these cells in bone does not fully mimic clinical human disease. It is well established that human PCa [[bone metastasis]] form osteoblastic lesions rather than osteolytic lesions seen in other cancers like breast cancer.<ref name="pmid10344214">{{cite journal |vauthors=Koeneman KS, Yeung F, Chung LW | title = Osteomimetic properties of prostate cancer cells: a hypothesis supporting the predilection of prostate cancer metastasis and growth in the bone environment | journal = Prostate | volume = 39 | issue = 4 | pages = 246–61 |date=June 1999 | pmid = 10344214 | doi = 10.1002/(SICI)1097-0045(19990601)39:4<246::AID-PROS5>3.0.CO;2-U | doi-access = free }}</ref><ref name="pmid11351351">{{cite journal |vauthors=Lin DL, Tarnowski CP, Zhang J, Dai J, Rohn E, Patel AH, Morris MD, Keller ET | title = Bone metastatic LNCaP-derivative C4-2B prostate cancer cell line mineralizes in vitro | journal = Prostate | volume = 47 | issue = 3 | pages = 212–21 |date=May 2001 | pmid = 11351351 | doi = 10.1002/pros.1065 | hdl = 2027.42/34759 | s2cid = 31076493 | url = https://deepblue.lib.umich.edu/bitstream/2027.42/34759/1/1065_ftp.pdf | hdl-access = free }}</ref> Similarly, PC-3 and DU145 cells form osteolytic tumors. To develop an AI-PCa cell model that more closely mimics clinical disease, LNCaP sublines have been generated to provide the most clinically relevant tissue culture tools to date.
==History==
The LNCaP ('''L'''ymph '''N'''ode '''Ca'''rcinoma of the '''P'''rostate) [[cell line]] was established from a metastatic lesion of human prostatic adenocarcinoma. The LNCaP cells grow readily in vitro (up to 8 x 10<sup>5</sup> cells/sq cm; doubling time, 60 hr), form clones and are highly resistant to human fibroblast [[interferon]].<ref name="pmid6831420" /> LNCaP cells have a modal chromosome number of 76 to 91, indicative of a human male karyotype with several marker [[chromosome]]s.<ref name="pmid6831420" /> The malignant properties of LNCaP cells are maintained in athymic nude mice which develop tumors at the injection site and show a similar doubling time ''in vivo''.<ref name="pmid6831420" />
High-affinity specific [[androgen]] and [[estrogen]] receptors are present in the cytosol and nuclear fractions.<ref name="pmid6831420" /> The LNCaP line is [[hormone|hormonally]] responsive, shown by ''in vitro'' 5 alpha-dihydrotestosterone modulation of cell growth and acid phosphatase production.<ref name="pmid6831420" /> LNCaP cells also express [[Prostate Specific Antigen]] (PSA).<ref name="pmid6831420" /> ''In vivo'', Male mice develop tumors earlier and at a greater frequency than do females and hormonal manipulations show that the frequency of tumor development correlates with serum androgen levels.<ref name="pmid6831420" /> The rate of the tumor growth, however, is independent of the gender or hormonal status of the host.<ref name="pmid6831420" />
==C4/C5 and C4-2==
<!-- Deleted image removed: [[File:Thalmann figure.png|thumb|Generation of LNCaP sublines.<ref name="pmid8168083">{{cite journal |vauthors=Thalmann GN, Anezinis PE, Chang SM, Zhau HE, Kim EE, Hopwood VL, Pathak S, von Eschenbach AC, Chung LW | title = Androgen-independent cancer progression and bone metastasis in the LNCaP model of human prostate cancer | journal = Cancer Res | volume = 54 | issue = 10 | pages = 2577–81 |date=May 1994 | pmid = 8168083 }}</ref>
]] -->
Wu et al. (1994) reproduced the human-derived LNCaP tumors in immunocompromised mice by co-injection of LNCaP cells with MS human bone fibroblasts.<ref name="pmid8169003">{{cite journal |vauthors=Wu HC, Hsieh JT, Gleave ME, Brown NM, Pathak S, Chung LW | title = Derivation of androgen-independent human LNCaP prostatic cancer cell sublines: role of bone stromal cells | journal = Int J Cancer | volume = 57 | issue = 3 | pages = 406–12 |date=May 1994 | pmid = 8169003 | doi = 10.1002/ijc.2910570319| s2cid = 22056818 }}</ref> Cells were subcutaneously injected at multiple sites into the mouse flank and after approximately 4 weeks of growth, tumors were easily detectable by physical examination and had a high rate of growth (17-33 mm3/day).<ref name="pmid8169003"/>
To replicate the hallmark shift of PCa cells to AI, LNCaP host mice were castrated by way of midscrotal incision at approximately 8 weeks post injection. Tumors were maintained in castrated hosts for 4 to 5 weeks at which time remaining tumors were harvested. In total, two subsets of cells were collected from castrated hosts: C4 and C5, collected at 4 and 5 weeks respectively.<ref name="pmid8169003"/>
To further select for AI-PCa cells, the C4 subline was co-injected with MS human fibroblasts into a castrated host. The resulting tumors were isolated and an additional subline was generated, C4-2.<ref name="pmid8169003"/>
'''[[Karyotype]] comparisons''' indicate that
LNCaP cells grown in intact hosts (M subline) have a modal chromosomal distribution number of 83, C4 and C5 sublines with 85, and the C4-2 subline with 83.<ref name="pmid8169003"/>
To further verify that these cells were of human origin karyotype analysis determined that the parental LNCaP cells had 7 distinct marker chromosomes, with two copies of each. The M, C4, C5, and C4-2 sublines contained most of the marker chromosomes, with the M subline being most similar to the parental LNCaP cells. C4, C5 and C4-2 are only minutely distinct from LNCaP and the M subline with the addition of a marker chromosome resulting from a segment addition to chromosome 6. A [[Y chromosome]] is not present in most C4, C5 and C4-2 cells, suggesting major chromosomal alterations.<ref name="pmid8169003"/>
C4, C5, and C4-2 sublines grow well under identical tissue culture conditions as LNCaP with similar growth rates. Parental LNCaP, M, C4, and C5 subline have similar baseline gene expression levels of [[ornithine decarboxylase]] (ODC) and [[Prostate Specific Antigen]] (PSA) however, M, C4, and C5 sublines express 5-10X more PSA mRNA. M, C4, C5 and C4-2 also expressed reduced human [[androgen receptor]] mRNA as expected in AI cells.<ref name="pmid8169003"/>
'''Androgen Insensitivity'''
All sublines were treated with [[dihydrotestosterone]] (DHT), a high-affinity ligand for AR. DHT treatment elicited markedly reduced growth in C4 and C5 cells and no growth in C4-2 cells when compared to the high rate of growth seen in LNCaP cells, suggesting reduced androgen sensitivity in C4 and C5 and AI in C4-2 cells. Whole-cell AR assay also indicated that LNCaP cells have a much higher affinity form of AR (Kd = 159 pM) when compared to C4-2 (Kd = 267 pM).<ref name="pmid8169003"/>
'''Tumorigenicity'''
C4 and C5 sublines exhibit greatly increased tumorigenicity when injected in intact male mice, unlike parental LNCaP cells. C4 and C5 were also able to form highly vascularized carcinomas in castrated mice when co-injected with MS human fibroblasts. The C4-2 subline more readily forms tumors in intact hosts than C4 and C5 sublines and they are the only cells able to form tumors in castrated host without co-injection of MS human bone fibroblasts. These same C4-2 tumors stained for PSA and secreted high levels of PSA into the growth medium.<ref name="pmid8169003"/>
==C4-2B==
To generate a bone metastatic subline, C4-2 cells were orthotopically injected into castrated male mice. These cells formed large primary tumors of the prostate, lymph nodes, as well as osseus tumors. Isolation of these osseus tumors resulted in the C4-2B subline.
C4-2B cells were positive for PSA and [[cytokeratin 8]], confirming their prostatic origin. Most importantly, immunohistochemical staining of the C4-2B tumors were positive for osteoblast activity suggesting the induction of osteoblastic tumor formation mirroring the progression of human PCa.<ref name="pmid8168083">{{cite journal |vauthors=Thalmann GN, Anezinis PE, Chang SM, Zhau HE, Kim EE, Hopwood VL, Pathak S, von Eschenbach AC, Chung LW | title = Androgen-independent cancer progression and bone metastasis in the LNCaP model of human prostate cancer | journal = Cancer Res | volume = 54 | issue = 10 | pages = 2577–81 |date=May 1994 | pmid = 8168083 }}</ref>
When cultured in a “promineralization medium” that contains [[ascorbic acid]] (known to promote skeletal-like ECM formation in osteoblasts) and a source of phosphate (for [[hydroxyapatite]] formation), C4-2B cells produce and retain approximately 8x more mineralized calcium than parental LNCaP cells. C4-2B cells also express higher levels of [[osteoprotegerin]] (OPG), [[alkaline phosphatase]], [[bone sialoprotein]] (BSP), [[Osteocalcin]] (OCN), [[RANKL]], and [[Osteonectin]] (OSN) mRNA, all of which are highly expressed by [[osteoblasts]].
Osteoblast promoter activity is also higher in C4-2B cells when compared to LNCaP, as indicated by [[Cbfa1]] transcription factor expression. Concomitantly, [[BMP7]], a known inducer of Cbfa1, is also more highly expressed in C4-2B cells, further suggesting many osteoblast-like properties.<ref name="pmid11351351"/>
== Androgen Independent Variants ==
'''LN95''' cells were derived from LNCaP cultured in RPMI-1640 (without phenol red) with 10% charcoal stripped fetal bovine serum.<ref name="ReferenceA">{{cite journal|last1=Pflug|first1=BR|last2=Reiter|first2=RE|last3=Nelson|first3=JB|title=Caveolin expression is decreased following androgen deprivation in human prostate cancer cell lines.|journal=The Prostate|date=1 September 1999|volume=40|issue=4|pages=269–73|pmid=10420156|doi=10.1002/(sici)1097-0045(19990901)40:4<269::aid-pros9>3.0.co;2-6|s2cid=19432542 }}</ref> LN95 cells differ from parental LNCaP cells morphologically, with pronounced dendritic extensions, and molecularly, with [[Androgen receptor]] variant expression similar to that of AR-V7<sup>High</sup> [[VCaP]] cells.<ref name="ReferenceB">{{cite journal|last1=Liu|first1=LL|last2=Xie|first2=N|last3=Sun|first3=S|last4=Plymate|first4=S|last5=Mostaghel|first5=E|last6=Dong|first6=X|title=Mechanisms of the androgen receptor splicing in prostate cancer cells.|journal=Oncogene|date=12 June 2014|volume=33|issue=24|pages=3140–50|doi=10.1038/onc.2013.284|pmid=23851510|pmc=4553036}}</ref> Notably, LN95 cells are significantly more tumour initiating than their parental counterparts ''in vivo''.<ref name="ReferenceA"/>
'''LNCaP-AI''' (or LN-AI) cells were derived from 6-Month culture of LNCaP parental cells in charcoal stripped serum.<ref name="ReferenceC">{{cite journal|last1=Lu|first1=S|last2=Tsai|first2=SY|author3-link=Ming-Jer Tsai|last3=Tsai|first3=MJ|title=Molecular mechanisms of androgen-independent growth of human prostate cancer LNCaP-AI cells.|journal=Endocrinology|date=November 1999|volume=140|issue=11|pages=5054–9|doi=10.1210/endo.140.11.7086|pmid=10537131|doi-access=free}}</ref> While LNCaP-AI are wholly androgen independent, they retain high expression of [[Androgen receptor]], low expression of AR-V7, and remain androgen responsive.<ref name="ReferenceB"/> The mechanism of the highly proliferative phenotype of LNCaP-AI cells appears to be loss of cell cycle regulator expression (p21, p16) and increased anti-apoptotic [[Bcl2]] expression.<ref name="ReferenceC"/><ref>{{cite journal|last1=Sampson|first1=N|last2=Neuwirt|first2=H|last3=Puhr|first3=M|last4=Klocker|first4=H|last5=Eder|first5=IE|title=In vitro model systems to study androgen receptor signaling in prostate cancer.|journal=Endocrine-Related Cancer|date=April 2013|volume=20|issue=2|pages=R49–64|doi=10.1530/ERC-12-0401|pmid=23447570|doi-access=free}}</ref>
== References ==
{{Reflist | 2}}
==Further reading==
* {{cite journal |vauthors=Li H, Lovci MT, Kwon YS, Rosenfeld MG, Fu XD, Yeo GW |title=Determination of tag density required for digital transcriptome analysis: application to an androgen-sensitive prostate cancer model |journal=Proc Natl Acad Sci USA |volume=105 |issue= 51 |pages= 20179–84 |year= 2008 |pmid= 19088194 | doi=10.1073/pnas.0807121105 | pmc=2603435 |bibcode=2008PNAS..10520179L |doi-access=free }}
==External links==
* [http://cell-lines.toku-e.com/Cell-Lines_3599.html Transfection data on LNCaP Cells]
* [https://www.cellosaurus.org/CVCL_0395 Cellosaurus entry for LNCaP]
* [https://www.cellosaurus.org/CVCL_4783 Cellosaurus entry for C4]
* [https://www.cellosaurus.org/CVCL_4782 Cellosaurus entry for C4-2]
* [https://www.cellosaurus.org/CVCL_4784 Cellosaurus entry for C4-2B]
* [https://www.cellosaurus.org/CVCL_M132 Cellosaurus entry for C5]
{{DEFAULTSORT:Lncap}}
[[Category:Human cell lines]]
[[Category:Cancer research]]
[[Category:Prostate cancer]]
| |||