I. HMEC Types that We Distribute

 

Cells are sent frozen (in dry ice) in ampoules containing 5 x 10-5 or 1 x 10-6 cells. Under special circumstances, we can arrange to send growing cells in flasks.  See "REVIEW section VIII." for more information on cell shipments.

 

A. FINITE LIFESPAN HMEC (untreated) (Chart 2)(section II.)

 

1. Post-selection (p16 non-expressing) reduction mammoplasty derived HMEC grown in serum free (MCDB 170 like) medium

 

We have large batches of post-selection HMEC frozen at around passages 7-10 that we distribute.  Depending upon the individual, these cells senesce around passages 14-25 (about 3 PD per passage)(see section II.C. Figure 1).  These cells are routinely available from women of different ages.  I most commonly distribute cells from specimens 184, 48R, 239 and 161.

 

2. Pre-stasis reduction mammoplasty derived HMEC grown in a serum containing (MM like) medium

 

Because pre-stasis cells exhibit only ~15-30 PD total, we have only limited quantities of cells available for distribution for specific requests.  Cells are supplied at passages 2 or 3; most growth ceases around passages 4-5.  Pre-stasis HMEC contain a greater range of phenotypes than the post-selection MCDB 170 grown cells. I most commonly distribute cells from specimens 184, 48, and 161.

 

3. Cells from non reduction mammoplasty tissues (i.e., mastectomies, benign tumors, gynecomastias)

 

Limited quantities are available. Cells grown from tumor tissues in MCDB 170 are not reflective of most tumor cells. Talk with me directly about these cells.

 

B. CARCINOGEN EXPOSED EXTENDED LIFE CULTURES (Chart 2)(section III.A. and figure 5)

 

Very limited quantities of benzo(a)pyrene treated specimen 184 extended life cultures, including the extended life precursors of the lines 184A1 (184Aa) and 184B5 (184Be), are available. Talk with me directly about these cells.

 

C. FINITE LIFESPAN HUMAN MAMMARY FIBROBLAST CELLS (section I.)

 

We have available for distribution stocks of fibroblast cells from several reduction mammoplasty specimens for which HMEC are available, particularly specimens 184, 48, and 161. These cells are grown in a serum containing medium.  In theory, fibroblast stocks can be obtained from any of our specimens, including the mastectomy derived tissues, but we have not grown up stocks to distribute from more than a few.  Frozen cells are available around passages 4-6, and they senesce around passages 10-20 (2-3 population doublings per passage) depending upon the individual.

 

D. PRIMARY TISSUES (section I.)

 

We have very limited quantities of frozen organoids, and are therefore very reluctant to distribute any of this material, but will make exceptions for specific studies. More primary tissue is available from reduction mammoplasties than mastectomies. Talk with me directly about these cells.

 

D. IMMORTALLY TRANSFORMED CELL LINES

 

1. Immortal lines derived following exposure to benzo(a)pyrene (p53+) (Chart 3) (section III.B-D.)

 

184A1

 

Unlimited quantities of later passages of this cell line of indefinite lifespan are available for distribution.  Very limited quantities of early passage conditionally immortal cells are available upon specific request.  Clonal isolates are also available.  These cells are wild type for p53, and RB, and are not anchorage independent or tumorigenic.  They have the most stable karyotype of our immortally transformed lines (not considering the hTERT immortalized lines).  This line is derived from the extended life precursor 184Aa.

 

More limited quantities of cells are available which have been selected for loss of specific nutritional requirements. (section III.C.1.)

 

We also have a lineage of fully immortal 184A1 defective in several p53 functions.

 

184B5

 

Unlimited quantities of later passages of this cell line of indefinite lifespan are available for distribution.  More limited quantities of early passage conditionally immortal cells undergoing conversion are available upon specific request.  Clonal isolates are also available. These cells are wild type for p53, and RB, and are not anchorage independent or tumorigenic.  They have a low level of karyotypic instability. This line is derived from the extended life precursor 184Be.

 

More limited quantities of cells are available which have been selected for loss of specific nutritional requirements. (section III.C.1.)

 

We have also transfected 184B5 with the breast cancer associated oncogene erbB2, producing B5-erbB2 and B5Me (isolated from anchorage independent colonies).  These cells now exhibit anchorage independent growth and a may have a very low level of tumorigenicity.  Unlimited quantities are available. (section III.C.2.)

 

184AA4

 

Unlimited quantities of later passages of this cell line of indefinite lifespan are available for distribution.  Early passage cells are not available. These cells are wild type for p53, and RB, and are not anchorage independent or tumorigenic.  They have many karyotypic abnormalities.  This line is derived from the extended life precursor 184Aa.

 

2. Immortal lines lacking p53 function (Chart 3) (section IV.C.)

 

184AA2

 

Unlimited quantities of this p53(-/-) cell line of indefinite lifespan are available for distribution. These cells are wild type for RB, are anchorage independent, and have some tumorigenicity.  The karyotype is unstable.  184AA2 is derived from the same extended life precursor population (184Aa) as 184A1 and 184AA4.

 

184AA3

 

Unlimited quantities of this p53(-/-) cell line of indefinite lifespan are available for distribution.   These cells are wild type for RB, are anchorage independent by passage 50, and have some tumorigenicity.  The karyotype is unstable.  They may be less deranged than 184AA2.  184AA3 is derived from the same extended life precursor population (184Aa) as 184A1 and 184AA4.

 

184A1-GSE22

 

We have transduced 184A1 at passage 12 (when still conditionally immortal) with a genetic suppressor element (GSE) that produces a peptide that interferes with p53 function.  These cultures provide a matched set of 184A1 with and without p53 function.  However, when the GSE22 is introduced pre-conversion, the process of conversion is altered.  An even more matched set would require transduction of the GSE22 into later passage fully immortal 184A1.  

 

184Aa-GSE22

 

We have transduced finite lifespan EL 184Aa with the GSE22 and produced immortally transformed cultures.  Immortalization is sporadic, and each isolate is likely to be somewhat different.  We presume these lines have acquired additional derangements during the period of agonescence/crisis. 

 

3. Immortal lines following exposure to potential breast cancer oncogenes (with and without exposure to benzo(a)pyrene) (Chart 4) (section IV.B.)

 

184-ZN4; 184ZN5; 184ZN6

184Aa-ZN1-3

 

Several different batches of post-selection184 HMEC and EL 184Aa have been transduced with the potential breast cancer oncogene ZNF217.  Immortalization is sporadic, and each isolate is likely to be somewhat different.  We presume these lines have acquired additional derangements during the period of agonescence.

 

184-myc

184Aa-myc

 

We also have post-selection 184 and 48R HMEC transduced with the p53 inhibitory genetic suppressor element GSE22.

 

These immortal lines are still under experimental observation. Talk with me directly about these cells.

 

4. Immortal lines following exposure to viral oncogenes (with and without exposure to benzo(a)pyrene) (section IV.E.)

 

184-E6

 

Transduction of the HPV16 E6 gene into post-selection 184 HMEC produces efficient immortal transformation.  Talk with me directly about these cells.

 

184A1- E6; -E7; -T; -E1A

 

Early passage (p12) conditionally immortal 184A1 has been transduced with either the HPV16 -E6, -E7, SV40T, or E1A genes.  These viral oncogenes produce numerous effects that may circumvent or alter the conversion process.  Talk with me directly about these cells.

 

5. Immortal lines following transduction of hTERT (Chart 4)(section IV.D.)

 

184-; 48R-; 161-hTERT

 

Finite lifespan post-selection HMEC are efficiently immortalized with hTERT without undergoing agonescence or conversion.  These cells are available for use when experiments require direct comparison to other 184, 48R, or 161 HMEC, and require permission from Geron Corp (more information pending). 

 

184A1-hTERT(12p)

 

Early passage (passage 12) conditionally immortal 184A1 was transduced with hTERT, resulting in immediate uniform immortalization without undergoing conversion.  These cells also require permission from Geron Corp (more information pending).

 

184A1-hTERT(22p)

 

Conditionally immortal 184A1 already in the process of conversion (passage 22) was transduced with hTERT.  These cells proceeded through conversion to become fully immortal.  These cells also require permission from Geron Corp (more information pending).