internal,medicine,RA,biotechnoogy,sepsis,airways and lungs,targeted therapy

1. Principles of “Targeted Therapy”

2. • Thanks to recent acquisitions in the field of molecular
biology it has been possible to identify new biological
targets expressed by tumor cells
• Using drugs directed against these targets it is possible
to hit, in a “targeted way” these cells
• One of the most studied target is the Epidermal Growth
Factor Receptor (EGFR)
TARGETED-THERAPY

3. • Signal transduction inhibitors
• Proteasomes inhibitors (Multiple Myeloma)
• Cox-2 inhibitors (Colon Poliposis)
• Vaccines, immunotherapy and gene therapy
TARGETED THERAPY IN CANCER

4. Nat Rev Cancer. 2009 Jan;9(1):28-39.
Targeting cancer with small molecule kinase inhibitors.
Zhang J, Yang PL, Gray NS.
Deregulation of kinase activity has emerged as a major mechanism by which cancer
cells evade normal physiological constraints on growth and survival. To date, 11
kinase inhibitors have received US Food and Drug Administration approval as
cancer treatments, and there are considerable efforts to develop selective small
molecule inhibitors for a host of other kinases that are implicated in cancer and other
diseases. Herein we discuss the current challenges in the field, such as designing
selective inhibitors and developing strategies to overcome resistance mutations. This
Review provides a broad overview of some of the approaches currently used to
discover and characterize new kinase inhibitors.
FURTHER READING

5. MetastasisProliferation
Cell-cycle progression
pY
pY
K K
pY
M
G1S
G2
Growth factor
SIGNAL TRANSDUCTION
INHIBITORS

6. • EGFR Inhibitors
– Gefitinib (Iressa)
– Erlotinib (Tarceva)
• EGFR Monoclonal antibodies
– Cetuximab (Erbitux)
• VEGF Monoclonal antibodies
– Bevacizumab (Avastin)
TARGETED THERAPIES

7. EGF VEGFBevacizumab
Gefitinib
Erlonitib
AZD
(TK inhibitor)
Cetuximab

8. myc cyclin D1
Jun Fos
RR
RR
RR
KK
K
Gene transcription
RR
K
EGFR SIGNALING

9. myc cyclin D1
Jun Fos
RRRR
KRR
K
RR
K
MAPKMAPK
MEKMEK
RASRAS RAFRAF
SOSSOS
GRB2GRB2
PI3-KPI3-K
AKTAKT
Proliferation
Inhibition of
apoptosis
K
PPPP
PP
PP
PP
PP
MetastasisAngiogenesis
GEFITINIB
Gene transcription
EGFR SIGNALING

10. Iressa received accelerated FDA approval in
May 2003 as a 3rd line therapy in NSCLC
based on the IDEAL-2 study.
GEFITINIB (IRESSA®
)

11. 0
10
20
30
40
50
60
70
80
90
100
Pazienti
Percentualedirisposta
Totale
Donne con
adenocarcinoma
non fumatrici
0
10
20
30
40
50
60
70
80
90
100
Pazienti
Percentualedirisposta
Uomini
Donne
Adenocarcinoma
Non fumatori
IRESSA®
: results in different subgroups of
patients with NSLC

12. pre-treatment post-treatment
RESULTS (I)

13. pre-treatment post-treatment
RESULTS (II)

14. • The efficacy of GEFITINIB is limited to a subset of
patients (nonsmoker women affected by adenocarcinoma)
• The overall response rate is 7%
• In nonsmoker women with adenocarcinoma the response
rate reaches 50%
• Gefitinib toxicity is lower with respect to that observed
with conventional chemotherapy
CONCLUSIONS

15. myc cyclin D1
Jun Fos
RRRR
KRR
K
RR
K
MAPKMAPK
MEKMEK
RASRAS RAFRAF
SOSSOS
GRB2GRB2
PI3-KPI3-K
AKTAKT
Proliferation
Inhibition of
apoptosis
K
PPPP
PP
PP
PP
PP
MetastasisAngiogenesis
ERLOTINIB
Gene transcription
EGFR SIGNALING

16. Tarceva
(n=488)
Placebo
(n=243)
Resp. Rate 8.9% <1%
Median
survival
6.7 mo 4.7 mo
PFS* 2.2 mo 1.8 mo
Shepherd FA et al. NEJM 2005
Most common AE: rash, diarrhea
*sopravvivenza libera da progressione
ERLOTINIB (TARCEVA®
) IN
NSLC
Tarceva received FDA approval in 2004 as a 2nd or
3rd line therapy in NSCLC based on Shepherd’s trial.
* Progression Free Survaival

17. • TALENT trial - Cisplatin/gemcitabine
+ (tarceva or placebo) - Phase III
– 1172 pts., previously untreated
Tarceva Placebo
Med. survival
(days)
301 309
TTP* (days) 167 179
Gatzemeier U. ASCO 2004 Abstract
*intervallo per ripresa di progressione tumorale
* Time To Progression
ERLOTINIB (TARCEVA®
) IN
NSLC

18. Group RR (%) p
Women v men 19 v 3 0.001
Japanese v
non-Japanese
27.5 v 10.4 0.0023
Adenocarcinoma v
others
13 v 4 0.046
BAC* v adeno 38 v 14 <0.001
Never smoker v
current/former
36 v 8 <0.001
Fukuoka JCO 2003;21:2237-46. Kris JAMA 2003;290:2149-58. Miller JCO 2004;22:1103-09.
* Bronchioloalveolar carcinoma
PREDICTORS OF RESPONSE TO
TK INHIBITORS IN NSCLC

20. Pao et al. JCO 2005; 23:2556-2568.

21. Some patients have significant initial
clinical response to TK Inhibitors, but then
develop progressive disease
– Secondary mutation found (4/7 pts.)
• Substitution of methionine for threonine at position
790
• Get steric hindrance of TK Inhibitor binding
Pao et al. 2005; PLoS Med 2:1-11.
Kobayashi et al. 2005; NEJM 352:786-92.
ACQUIRED RESISTANCE TO TK
INHIBITORS

22. OTHER MUTATIONS
Pao et al. JCO 2005; 23:2556-2568.
Mutation

23. • Mutations in K-ras
– Occur in 15-30% of lung adenocarcinomas
– High incidence in heavy smokers
• Predict a poor response to TK inhibitors
• EGFR and K-ras mutations are mutually
exclusive
Ahrendt SA et al. Cancer 2001; 92:1525-30.
Kosaka T et al. Cancer Res 2004; 64: 8919-23.
Eberhard DA et al. JCO 2005;23:5900-09.
MUTATIONS IN K-ras

24. • Many centers have started testing for the
most common mutations in EGFR
• Problem:
• What to do if negative for mutations?
– Some pts. with responses to Gefitinib/Erlotinib have no
detectable EGFR mutation
MUTATIONAL PROFILING

25. ANTI EGFR MONOCLONAL
ANTIBODIES
Pao et al. JCO 2005; 23:2556-2568.
mAb
(CETUXIMAB)

26. RR (%)
(CI)
Median
survival
(mo.)
Median
TTP*
(mo.)
Cisplatin/Vinor.
20
(7.6-32.4)
7.0 4.2
Cisplatin/Vinor.
+Erbitux
32
(17.5-46.0)
8.3 4.7
Rosell 2004 ASCO Abstract
CETUXIMAB WITH CHEMO:
RESULTS
RR: response rate
TTP: time to progression

27. Survival
in circulationIntravasation
Invasion
initiation
of growth
Sustained
growth
Primary
tumour
Clinically
evident metastasis
Adapted from Chambers A, Matrisian L. J Natl Cancer Inst 1997;89:1260–70
ANGIOGENESIS:
MULTIPLE STEPS IN CANCER BIOLOGY

28. Sustained Growth
requires blood vessels
Primary
tumor
Metastasis
VEGF
EGF
Multiple steps in cancer biology
Heading towards Personalized Care

29. ANGIOGENESIS

30. ANGIOGENESIS IN TUMORS

31. Endostatin is a
natural
antiangiogenic
protein that
inhibits the
growth of blood
vessels
Vaccines targeting tumor angiogenesis—a novel strategy for cancer immunotherapy Y. Okaji et al, 2006
ANGIOGENESIS INHIBITION

32. Nature Reviews Drug Discovery 4, 448-449 (2005)
Anti-angiogenesi (II)
Bevacizumab (Avastin, Genetech):
Vascular Endothelial Growth
factor (VEGF) inhibitor.
Phase III Trial in NSCLC:
- Survival of 12.5 months in
Avastatina/CHT arm vs. 10.5
months in conventional CHT arm
- 2 years survival: 22% vs. 17%
- Adverse events: 5% major
bleedings, 1% mortality

33. Avastin + Chemo
Carbo/Taxo
l
(n=32)
Carbo/Taxol
+ Avastin
(n=35)
Median
survival
(mo.)
14.9 17.7 (NS)
RR (%) 18.8 31.5
TTP* (mo.) 4.2 7.4
Johnson et al., JCO 2004.Johnson et al., JCO 2004.(19/32 pts. crossed over)
Avastin in combination with Tarceva appears to have a
synergistic effect
*intervallo per ripresa di progressione tumorale

34. Animal Model to study angiogenesis

35. Growth Factor Reduced BD Matrigel Matrix
Storage -30°C
Melting +4°C
Solidification +37°C
Matrigel is an extracellular matrix produced by murine
sarcoma Engelbreth-Holm-Swarm (EHS), consisting
predominantly of laminin (60%), collagen IV (30%),heparan
sulphate (5%), proteoglycans (3 %) and entactin (1%).

36. • Matrigel subcutaneous injection (the matrix can be
mixed with different molecules to be studied).
• After the injection, thanks to body temperature,
the matrix solidifies and leads to the formation of
“plugs”. With appropriate stimuli the angiogenic
process starts and develops into the plugs.
Model for in vivo study of angiogenesis using
Matrigel Matrix

37. MATRIGEL PLUGS

38. Reperimento dei plugs  dopo 7 giorni dall’inoculo
 sezione addominale lungo la linea mediana
 isolamento masserelle dai tessuti
 conservazione del materiale in apposite biocassette
Allestimento sezioni istologiche  fissazione in formalina
 inclusione in paraffina
 taglio con microtomo
 allestimento vetrino
 colorazione

39. RESULTS EVALUATION

40. NEGATIVE CONTROLS
MATRIGEL MATRIGEL + Gas 6