1. Eternal Life: Cell Immortalization and Tumorigenesis
Growth Patterns and Pathobiology of Experimental and Clinical Malignancies In Vivo
Folder title: Growth(NoTP)
Chapter 10, 2nd Edition:
Eternal Life: Cell Immortalization and Tumorigenesis pp
Folder Title: Growth
Neoplasia is by definition, growth of aberrant cells. Therefore, the basic biology underlying the growth of cancer cells and of organized tumor masses is crucial to understanding Cancer Biology and to attempts at cancer management based on controlling aberrant cell growth. Most clinically effective anti-neoplastic protocols involve the use of agents that affect cell growth.
Updated: March 20, 2017

2. Pre-Clinical Breast Carcinoma (from Gullino, Cancer, 39:2697-2703)
103

3. Questions About Neoplastic Cell Growth
How do we measure neoplastic cell growth?
In Vitro in cell culture or in tissue culture?
In vivo in animals?
In vivo in patients?
What are cancer growth patterns like?
What causes the patterns of growth that we see?
What maintains those growth patterns?
Can we alter those patterns to benefit patients?

4. Measurement of Cancer Growth
In Cell Culture
Cell numbers vs time
Colony formation assays (See next slide for cloning assays)
Labelled thymidine incorporation
Dye reduction assays for viable cells (Dye reduction by mitochondrial activity)
Dye exclusion assays by viable cells (Dye exclusion by intact plasma membranes)
In Vivo in Experimental Animals
Time to appearance of measurable tumor nodules
Dimensions of tumor nodules with time
Measurement of tumor products (e.g tumor antigens)
Host survival time
(Some guidelines on experimentation with live animal models, survival assays, and compassionate euthanasia)

5. Human U937 Leukemia Cells
Cloned in Soft Agar
200 Cells Cloned
Per Tube.
Stained with
pINT (tetrazolium dye).
Clones counted:
106
110
Cloning Efficiency: 53% %

6. Measuring Tumor Growth in Vivo in Patients
Brain Cancer Scans

9. Pathological Effects of Cancer on the Host
Inflammation: Macrophage, Granulocyte Infiltration, Cytokine Production
Cachexia (Wasting)
Hemorrhaging and Clotting
Immunosuppression
Infection
Pathological Production of Hormones and Other Bio-active Agents
Loss of Essential Functions: Tissue Destruction and Vital Organ Failure

10. Proliferation Control Differences: Normal Cells vs Neoplastic Cells in Cell Culture
Density-dependent Inhibition of Mitosis:
(Contact Inhibition of Mitosis)
Normal Fibroblasts: 40,000 cells/cm2
Virally Transformed Fibroblasts: 4 million cells/cm2
Anchorage-Dependent Effects on Proliferation
Normal Cells
Grow Attached and Flattened
Stop Dividing When Rounded by Contact
Stop Dividing if Artificially Rounded even with no Contact
Restart Dividing when monolayer is "wounded"
Transformed Cells
Keep Growing when rounded by contact (overgrow)
Keep growing if artificially rounded even wth no contact Will form colonies in viscous (semi-solid medium)
Note key role of cytoskeleton in cancer cell growth in vitro and in vivo

11. Control of Tumor Growth Rate In Vivo: Consequences for Concepts of Cancer Treatment
Concepts Involving Chemotherapy
Use and limitations of cell-cycle specific agents
Re-seeding of tumor growth by non-cycling cells
Concepts Involving Tumor Progression
Cell Loss and Cell Selection
A Microcosm of Evolution
Options for In Vivo Therapeutic Control
Anti-angiogenesis
Promotion of apoptosis (programmed cell death)
Exploitation of tumor micro-environment: pH
Tumor necrosis factor
Combination chemotherapy
Cell Synchronization and Cell-cycle-specific treatments
Host response modification
Induction of Differentiation
Targeting the cancer cell phenotype: e.g cell size and cell division

12. Managing Cancer Development, Progression and Growth In Patients
Inhibit Cell Production!
Promote Cell Loss!
Drive out of the Cell Cycle
Induce differentiation toward normal
Block host support

13. Control of Tumor Growth Rate In Vivo: Cell Production
Cell cycle Time
Growth Fraction: Proportion of Cells in the Cell Cycle to those in Go (not cycling)
Proportion of Clonogenic Stem Cells
Clonogenic Stem Cells
Probability of cell renewal:
0.5 = No net growth
1.0 = 100 billion cells after 36 divisions
As for bacteria
No cell death
No cell differentiation
0.51 = 100 billion cells after 1500 divisions
inexorable, slow, relentless growth vs rapid exponential growth
implications for therapy

14. Control of Tumor Growth Rate In Vivo: Inhibit Aberrant Cell Production
Blocking hormone access for hormone-dependent cancers.
Induction of differentiation with cytokines or other signals.
Block aberrant cell signaling pathways
Block Host Response Support: Anti-angiogenesis
Block DNA Repair Pathways
Show Movies from Biology of Cancer DVD
8.1 Animal Cell division
3.1 Contact Inhibition
2.2 Mammary Cancer Cells 2.3 Lymphoma

15. Control of Tumor Growth Rate In Vivo: Cell Loss
Cellular Anaplasia
Disrupted Cytoskeletal Structures
Aberrant Mitoses
Nuclear Abnormalities
Positional Anaplasia
Poor Blood Supply; Aberrant Vasculature
Low Oxygen Tension, Acidic pH
Host anti-Tumor Responses
Hormonal Effects
Immunological Effects
Retained Ability to Respond to Normal Signals
Apoptosis: Genetically Programmed Cell Death
Retained Recognition of Cell Senescence Programs
Applications in Chemotherapy, Radiation Therapy, and Immunotherapy

16. Monitoring Growth in the Therapy of Mammary Carcinomas Lacking BRCA2 Mediated DNA-Repair Capacity

17. (See Figure 12-40 and Sidebar 12.11, p. 520 Weinberg)
Chemotherapy Directed Toward Defective BRCA-1 and BRCA-2 Genes in Breast and Ovarian Cancers
(See Figure and Sidebar 12.11, p. 520 Weinberg)
Propositions:
Redundant DNA-repair mechanisms needed by both normal and neoplastic cells to repair DNA lesions incurred normally during cell division.
Repair of damaged DNA is even more important if chemotherapy with DNA-directed anticancer agents or radiation therapy is being carried out.
One type of DNA repair involves poly-ADP-ribose polymerase (PARP).
BRCA-1 and BRCA-2 have DNA repair functions as “Housekeeping genes”.
Normal cells can use BRCA-1 and BRCA-2 repair functions as well as PARP repair mechanisms.
Breast and Ovarian cancer cells lacking BRCA-1 and BRCA-2 must rely on the PARP repair option.
What happens if one inhibits the PARP repair function using PARP inhibitors, especially during treatment with agents damaging DNA?
Filename: BRCA2Therapy.ppt

18. Figure Weinberg
Cells lacking BRCA-2 (red line) are killed off (2 log kill or 99% kill) at 10E-7 M (0.1 uM) anti-PARP drug concentration. Anti-PARP prevents DNA repair. (The structure of the inhibitor is not specified).
Cells with BRCA-2 repair (blue or green lines) can survive almost 1,000-fold higher concentration of anti-PARP agent (10E-4M or 100 uM) because they can use BRCA-2 for DNA repair.
If both repair options are blocked the cancer cells die at low drug concentration of anti-PARP agent
PARP = poly-ADP-ribose DNA repair enzyme
BRCA2 = breast cancer associated antigen involved in DNA damage repair.

19. Three Turning Point Questions.
Please put away all other stuff.

22. Response Counter

23. Response Counter

24. Senescence, Aging, and Immortalization in Normal and Cancer Cells:
Figures from Biology of Cancer: Second Edition, Chapter 10

25. Senescence of Normal Cells (Human Fibroblasts) in Culture

26. Senescence of Cells In Cell culture
Senescent
Pre-senescent

27. Ageing in dermal fibroblasts

28. Apoptosis and death of normal cells.
Cell fragments (Apoptotic Bodies)
Intact
Cell
Apoptosis and death of normal cells.

29. Telomerase, Resynthesis of Chromosomal Telomeres, and Cellular Senescence

30. Preferential Destruction of Leukemia Cells Based on Size:
Untreated Human Promyleocytic Leukemia Cells vs
Cells Treated with Microfilament-Directed Agent (Cytochalasin B) to Produce Leukemia-specific Cell Enlargement
Application of Colony-Formation Assay in Monitoring Effects of Chemotherapy-Physicotherapy
Of Leukemia Cells in Cell Culture

31. Untreated U937 cells are reduced in cloning efficiency by less than 20% when sonicated for 4 minutes at 20 watts
Seeded 200 Cells

32. Drug Treated
Drug Treated and Sonic Treated
Seeded 400 Cells
Cloning efficiency of U937 cells treated for 12 hours with 2 uM CB is cut by 60% with sonication for 4 minutes. This compares to 20% reduction in clonogenicity of untreated U937 cells or 40% reduction with drug-treatment alone.

33. Some Thoughts on Physical Approaches to the Management of Cancers:
Leukemias and Blood-borne of Lymphatic Fluid Metastases
Application from outside of the patient
Precise control of dosage
Opportunity for repetitive treatment
Whole body application
Directed appendage application
Application to extra-corporeal shunt
Evasion of chemically-based drug resistance (unless combined with chemotherapy to alter size)

34. End Growth Presentation
Story of Hana Leon
Who is My Sister?

36. A visit to the place where Hana Leon, homeless, froze to death
The place where Hana Leon, 42, homeless, died on Feb. 27. She was found under the evergreen by a passerby. (Dennis Nett |
By Steve Carlic | on March 12, 2014 at 8:11 AM, updated March 17, 2014 at 9:33 AM
Hana Leon died in a place you would never want to.
She died on Feb. 27, beneath a small, ornamental evergreen, tucked behind a decorative brick wall that marks the entrance to Franklin Square. It's on the west side of Franklin Street, beside the on-ramp to S. West Street. There's a black and gold plaque attached to the brick that recognizes the 1990 National Economic Development Partnership Award given to the city of Syracuse and Pyramid Companies for restoring the neighborhood.
From where she lay, if you looked to the right you can see the old Learbury building, and ahead the brick facades of refurbished factories.
On Tuesday the only thing remaining at the spot were a dozen or so plastic bags stuffed with some socks, moldy food and handfuls of smaller plastic bags. It's not clear if these were Leon's worldly possessions or if they were dumped there since by somebody else.
Authorities say they think Leon, 42, died of hypothermia. She was found following a frigid night where the low was minus 8 degrees with the windchill.
Leon suffered from severe paranoid schizophrenia. She was among the 100 or so chronically homeless who live in Syracuse, where five homeless people have died over the past two winters.

37. Duke University Medical Center Dr. Henry Friedman
Glioblastoma and Glioblastoma Multiforme
Interveiwed by Dr. Sanjay Gupta
2010

38. Turning Point Questions
Please clear your desk of everything except your
NXT Transmitter

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