Day 4: Therapeutic strategies CRC, Targeted and Combination therapy Flashcards
HC 11, 12
HC11: Stage 0 CRC therapy
N0, M0: partial colectomy
> cancer cells are at mucosa or inner lining
> detected with colonoscopy
> mutation APC: adenomas
CRC stage 1 therapy
Local excision of polyp
> partial colectomy
> N0, M0
> cancer cell growth through mucosa and invade muscle layer
CRC stage 3 therapy
Partial colectomy, adjuvant chemotherapy (5-FU/LV, capecitabine) when high grade or spread
> N0, M0
> Cancer cells have grown to/through serosa and may spread to nearby organs
CRC stage 3 therapy
Partial colectomy, removal nearby LNs, adjuvant chemotherapy
> Cancer cells have grown through serosa
> spread can be observed to nearby organs
> cancer cells have spread to 1-6 LNs
> N1, M0
> adj chemo:
- 5-FU/LV/oxaliplatin (FOLFOX)
- capecitabine/oxaliplatin (CAPOX)
CRC stage 4 therapy
Partial colectomy, Neo/adjuvant chemotherapy
> N1/2, M1
> cancer cells have grown through serosa
> cancer cells may have spread to lymph node
> presence of distant metastasis (eg liver)
Chemo
- FOLFOX, FOLFIRI, CAPOX, FOLFOXIRI, FTD-TPI
Targeted
- VEGF inh
- EGFR inh
- immune checkpoint inhibitors
- Multi-kinase inh
5-FU chemotherapeutic
Will be transformed in compound that blocks Thymidylate synthase (by Tymidine phosphorylase and Thymidine kinase)
> TS blocking inhibits dTTP generation for DNA replication and repair: depletion by repressing TS > DNA damage > cell death
5-FU/LV:
LV (leucovorin) helps 5-FU to better bind the TS enzyme (Tymidylate synthase)
Capecitabine
Chemotherapeutic
> Precursor of 5-FU
> Same profile, survival, disease progression
> other side effects
> 5-FU has more stomatitis or neutropenia whule capecitabine will give more hand-foot syndrome
Platinum-based drugs
Chemotherapeutic
Like oxaliplatin
> hydrolysis to active drug
> Cisplatin binds N7 of guanine in C-G base pair (2x)
> forming intrastrain adducts: DNA damage
> apoptosis induced
Irinotecan (SN-38)
Chemotherapeutic
Transformation into SN-38 by a carboxylase
> SN-38 inhibits TOP1 (topoisomerase 1, for unwinding DNA for replication when replication fork formed)
> Collision with replication forks > double stranded DNA breaks
> DNA replication inhibited
Trifluridine (FTD/TFT)- Tipiracil (TPI)
Chemotherapeutic > third-line treatment
Analog of thymidine: but methyl group to CF3 group
> misincorporation of nucleoside analog (F3dTTP) into DNA
> DNA replication stress (DRS) and replication fork stalling
> lethal DNA damage
> kill the cancer cell
> TPI has antiangiogenic effect
Combining chemotherapeutics: FOLFOX
5-FU/LV + Oxaliplatin
> better survival rate than single agents
CAPOX (and vs FOLFOX)
Capecitabine + oxaliplatin
> overall survival equal efficacy
> in stage 3: CAPOX better than FOLFOX
> CAPOX is slightly more toxic than FOLFOX
> FOLFOX for 6 months and CAPOX for 3 months have same efficacy
FOLFIRI (and vs FOLFOX)
5-FU/LV/Irinotecan
> Similar outcome as FOLFOX
> different side effects
> FOLFOX: neurological problems
> FOLFIRI: heart rate decrease, hair loss
» primary goal is survival though, because side effects are bad
FOLFOX/FOLFIRI doublet vs FOLFOXIRI triplet
> more toxicity in triplets
more toxic products: more cancer cells and healthy cells affected
Which cells are affected by (systemic) chemotherapeutic treatments?
Any fast dividing cell
Chemotherapeutics generally cause …
DNA damage in replicating cells
> TS inhibition (5-FU)
> Adducts (platinum)
> TOP1 inhibition (irinotecan)
> misincorporation of nucleoside analog (FTD-TPI)
» accumulation DNA damage leads to cell death
Which healthy tissues are also affects by chemotherapy?
With quick turnover: bone marrow, digestive tract
> advanced CRC treatment: combining chemo is better but not without side effects
Targeted therapy concept
Identify important components of CRC signalling pathways to target it in specific manner
4 targeted routes CRC therapy
-VEGFR
-Wnt
-EGFR
-Immune checkpoint (CRC – T-cell)
Mutation in Wnt in cancer that forms target, and name Wnt characteristics
APC mutated
> Target: inhibit Wnt signaling
> APC part of b-catenin destruction complex
» proliferation and stemness
> Wnt also for tissue homeostasis in intestinal epithelial lining
> in bone: balance osteoblast and osteoclast (bone formation and resorption)
> in skin: homeostasis
Porcupine inhibitors
Inhibit Wnt secretion
Inhibit Frizzled of Wnt pathway
Antibodies
Tankyrase inhibitors
Block ubiquitination of Axin (part b-catenin destruction complex)
Problems with Frizzled antibodies, Porcupine inhibitors and tankyrase inhibitors for cancer
APC mutated: downstream b-catenin already stabilized, upstream targeting has no effect and no therapeutic benefit
Blocking Wnt with lncRNAs
- Restricted expression profiles in cell types: targeting specific tissues
- lncRNAs display high tissue specificity
- hit lncRNA in Wnt pathway are involved
> siRNAs, ASO, CRISPR to interfere with RNAs
Inducable shRNA against b-catenin
Tested in all CRC cell types known
> most CRC cells dependent on b-catenin: lowered cell proliferation
> some are independent or find bypasses
CRISPRi-based dropout screen
Epigenetic regulator is recruited: repress transcription at promotor
> repress expression of lncRNAs involved in cancer
> assess functionality of lncRNAs in cancer
CRL-1 as target
CRC specific expression
> ideal candidate
> b-catenin regulated lncRNA
> reactivated for proliferation in cancer
> cytoplasmic RNA
> use shRNA to block it
> CRL1 essential for CRC cell proliferation and tumor growth
CRL1 function (b-Catenin Regulated LncRNA-1)
CRL1 controls Myc protein levels
> CRL1 supports appropriate Myc and inhibits miRNA
> Myc and CRL1 both target genes of b-catenin
KO CRL1 in CRC
More differentiation, less stemness
> increase keratin
> decreased Lgr5 (marker stem cell)
> POLR1A upregulated in control: depletion marks terminal differentiation (in KO CRL1)
Steps towards RNAi based therapy against CRL1
1 siRNA screen
2 siRNA formulation
3 in vivo testing
> siRNA packaged in LNPs (lipid nanoparticles)
HC12: VEGFR and EGFR
Receptor Tyrosine Kinases
> to activate MAPK route
> mutated in patients with BRAF and KRAS mutations (oncogenic)
VEGF/VEGFR effect
Angiogenic effect on tumor
When hypoxia in tumor?
When oxygen demands cannot keep up with proliferation
> hypoxia and VEGF upregulate abnormal angiogenesis in cancer development
VEGF/VEGFR routes
Uses Akt-mTOR route and MAPK route for cell cycle activation and survival
Therapies against VEGF route
-Antibodies against VEGFR (eg bavacizumab)
-Protein kinase inhibitors (PKI) which inhibit tyrosine kinase intracellular part of VEGFR
Doublet bevacizumab + FOLFOXFIRI vs bevacizumab + FOLFIRI
Triplet has better survival but more side effects
> Adding bevacizumab also increases survival
When is blocking VEGF with bevacizumab efficient
In KRAS and BRAF WT patients, when mutated lowered effect
> Then, MAPK pathway independent of VEGFR signalling
Which mutation is worse: BRAF or KRAS
BRAF, further downstream in pathway, all upstream cannot be targeted for therapy
EGF/EGFR activated routes
The routes activated are
- PI3K to Akt
- Ras to Mek/Erk pathway (MAPK)
EGFR overexpression is associated with
- LN (lymph nodes affected)
> Tumor stage
> may promote more aggressive CRCs
Targeting EGFR
Cetuximab: mAb against EGFR
> block PI3K/Akt pathway (survival) and MAPK pathway (proliferation)
Which patients respond worse to cetuximab?
KRAS and BRAF mutant patients
> reactivation MAPK route independent of blocking EGFR
> resistance mechanism
Resistance mechanisms against chemotherapeutic cetuximab
- EGFR mutation
- Downstream mutations (KRAS, BRAF)
- Upregulation of other receptors
Methods to counteract resistance
- Change targeting strategy (to downstream)
- Broader approaches
- Combination therapy
Regulation interaction in KRAS and common mutation
P-loop in KRAS (GTP binding)
> G12 residue mutation in P-loop in KRAS: lock into active state
Inhibition KRAS mutant in mCRC G12C (common mutated KRAS)
Keep it into the GDP bound state specifically
Sotorasib / adagrasib
Makes covalent bond with KRAS (irreversible)
> selective for KRAS G12C
> Interact with inactive GDP bound form
MRTX1133
Make noncovalent bond with KRAS (reversible)
> selective for KRAS G12D
> Interacts with both active and inactive form (GDP)
How to improve adagrasib monotherapy
Adagrasib + cetuximab combination
> better PFS: progression free survival
> mono for better overall survival
Specific inhibition of BRAF mutants (V600E/D/K)
BRAFi
> Encorafenib: block catalytic activity (bind ATP pocket)
What drugs to use when BRAF mutation (7-10% of CRC)
MEKi
> Binimetinib or Trametinib: allosteric inhibitors (bind unphosphorylated Mek and prevents its phosphorylation)
Efficacy of BRAFi and MEKi and EGFRi compared to control of FOLFIRI and EGFRi
better survival
Expression Her2 in advanced CRC
Overexpression
> overexpressed in 3-5% of mCRC (metastasized)
Her2 in cancer
Induces genomic instability or amplification
> Her2 receptor
> Activation PI3K and MAPK pathway
Treating Her2 OE (overexpression)
- Trastuzumab: mAb against Her2 (receptor)
- Tucatinib (KI) against kinase if Her2 intracellularly
- Tucatinib + trastuzumab dual therapy: good response: can interfere with metastatic cancer
Regorafenib
Multi-kinase inhibitor
> inhibits intracellular kinases for eg VEGFR, FGFR, and more (but also of Raf!)
> block angiogenesis, cell proliferation, metastasis and immunosuppression simultaneously.
> to prevent resistance
> retains the dephosphorylated inactive state of kinases
TAS-102 as third line treatment
TAS-102 = Trifluridine-Tipiracil (FTD-TPI)
Broad anticancer treatment when refractory mCRC resistant against all therapies
> gain of 2 months
> improved when also adding bevacizumab
Why important to diversify and combine treatments
Adaptive rewiring of cancer cells via selection
> for example Mek1 mutation or NRAS mutation when therapies given
> block KRAS: cell uses more PI3K pathway
> cancer cells crosstalk: compensate to activate what the cell needs
Immune checkpoint: interactions CRC cell and T cell
Tumor cell – T-cell
B7 – CTLA4
MHC-I with peptide – TCR
PDL1 – PD1
> B7 and PDL1 give ‘don’t kill me’ signals to T-cell
Immune checkpoint inhibition
Antibodies against CTLA4, PD-1, PD-L1, B7
> reactivate the immune system against cancer cells
Why do a lot of mutations in the unstable genome of cancer benefit immune therapy?
More neo-antigens made by cancer cells: unusual proteins because mutations presented.
Mismatch repair mutated (MSI-H, dMMR) and immunotherapy
No repair in making neo-antigens > better efficacy immune therapy
Most CRC tumors and MSS/MSI
Most MSS or MSI-L (low), pMMR (proficient Mismatch Repair), less neo-antigens,
> less benefit from immune checkpoint inhibition
> more benefit when dMMR, MSI-H: more immunogenic
Which mutation makes patients beneficial for ICI (immune checkpoint inhibition)
PolE mutation (polymerase) > proofreading mutation > ultramutated genome even when MSS > higher neo-antigen presentation via MHC1 > more tumor-infiltrating lymphocytes and better prognosis
» via Nivolumab to inhibit MMR
Effect TGF-beta signalling on ICI response
Attenuates response: contributes to exclusion of T-cells
How to turn cold tumors hot
Inhibit TGFb with inhibitor and ICI
> block the dendritic cells from inhibiting T-cells
> immune attack once again: hot tumor
> first: T-cell response blocked: cold tumor
> only effective when dual therapy
Regorafenib effect on anti-tumor immunity
Improves it
> Normalize vasculature: facilitate access for CTLs
> Reduce amount of TAM/TEM (Tumor-associated macrophages and TIE-2 expressing monocytes): promotes T-cell activity
Potential resistance mechanisms against ICI
-Agonistic interaction: make T-cells better for attack
> antagonistic interaction should be blocked, could give rise to resistance
Oncolytic viruses therapy
- Healthy cells can better prevent a virus from dividing than cancer cell
- Cancer cells are less able to do this: cancer cells die, virus multiplies
> cell contents released: inflammation and destruction tumor microenvironment
Onyx-015
E1B deleted adenovirus
> P53 can not mitigate stress response > cell dies and virus replicates
> normally: p53 and Rb bind E1A and E1B respectively
Armed oncolytic viruses
Potentiating: put ICI encoded in viruses
> ONCR-177
> Transgenes delivered: activate T-cells, chemokine attractants made, expand and activate NKcells and DCs, ICI (anti-PD1 and anti-CTLA4)
Limitations oncolytic viruses
- Antiviral response > viral clearance
- Poor distribution throughout the tumor
> Tumor heterogeneity/ tropism (target cells p53 mutated, low IFN etc) - Dosing strategies
Adoptive Cell Transfer (ACT) therapies
Autologous cell therapy
> own cells cultured and given back
Allogenic cell therapy
> donor cells which are engineered and given to patient
CAR T-cell therapy
Chimeric antigen receptor (CAR) T-cells
> T-cells have group of proteins to make this receptor, but now chimeric protein made (all in one protein, easy to modify)
- IL-12 inducer intracellular: chemokine which helps activation T-cells
- IL-2Rb in next gen CAR: cytokine receptor (IL-2 helps T-cell proliferation, made upon binding ligand)
-Make CAR T-cells from T-cells and give back
> reognition of cancer specific epitope
Limitations CAR T-cells
-Infiltrating tumor mass (especially for poorly irrigated solid tumors)
-Immunosuppressive tumor microenvironment: hypoxia, high acidity, low concentration nutrients, TGFb
-On vs Off tumor toxicities:
> targeted antigens/ tumor specificity
> high cytokine production: aggressive immune reaction: tissue damage: organ failure
- Allogenic tranfer may be rejected by host (alloimmunization)
- autologous cell transfer: variability in starting material.
Best treatments for KRAS G12C mutant
Choose from
> anti-VEGFA: bevacizumab
> KRAS G12C inhibitor
> Regorafenib (against RTKs)
> Cetuximab (anti-EGFR)
> BRAFi and MEKi
BRAFi+MEKi
KRAS G12C inhibitor
Which patients benefit from ICI (choose one or more)
> MSS tumors
> POLE mutation: polymerase epsilon
> MSI-H
> low miRNA-200 expression
> CMS4 subtyped tumor with prominent activation of TGFb
> POLE mutants
MSI-H
