Chemotherapy Exam 2

Angiogenesis Inhibitor - Rationale

• tumors require new blood vessels (angiogenesis) to provide oxygen and nutrients for supporting growth beyond certain sizes

• blocking angiogenesis may block tumor growth

• Vascular endothelial growth factor (VEGF) plays a key role in tumor-induced angiogenesis

Bevacizumab (Avastin)

First-in-class VEGF inhibitor

Bevacizumab (Avastin) - Target

VEGF

• monoclonal antibody against VEGF

• VEGF is a ligand to VEGF receptor (VEGFR)

Bevacizumab (Avastin) - Mechanism 

• binds to VEGF (ligand) 

• prevents the ligands from binding to VEGFR 

• Blocks the activation of VEGFR

Bevacizumab (Avastin) - Clinical use

• in combination with chemotherapy for multiple cancers

  • lung, colon, renal, ovarian, brain cancers

• Modest effects

  • Ex: a median survival benefit of 4-5 months for patients with metastatic colorectal cancer (in combination with chemotherapy)

• Potential reasons for modest effects

  • hypoxic environment stimulates a compensatory VEGF production (feedback mechanism)

    • compromised blood supply causes a hypoxic (low-oxygen) environment

    • Hypoxic environment simulates the production of VEGF and other growth factors

  • Differential sensitivity of tumor blood vessels

  • Compromised vasculature reduces the efficiency of drug delivery to tumors

Bevacizumab (Avastin) - Side effects

hypertension, bleeding, impaired wound healing 

• VEGFR signaling affects nitric oxide synthesis (regulates blood pressure) and normal blood vessel survival and integrity 

Bevacizumab (Avastin) - Resistance

• increased level of VEGF

• Upregulation of other pro-angiogenic factors (ex: FGF) and receptor signaling

  • potential combating strategies: combining with other receptor tyrosine kinase inhibitors

Apoptosis inducer - Rationale

• Some cancer cells are particularly reliant on elevated levels of anti-apoptotic proteins to survive 

  • over-expression of anti-apoptotic proteins (ex: Bcl-2)

  • Alterations in cellular responses that increase reliance on anti-apoptotic proteins

• inhibiting anti-apoptotic proteins could restore and promote programmed cell death in cancer cells

Venetoclax (Venclexta)

First-in-class BCL-2 inhibitor

Venetoclax (Venclexta) - Target

BCL-2

• Bcl-2 is an anti-apoptotic protein

• Bcl-2 sequesters pro-apoptotic proteins (Bax and Bak), preventing them from forming dimers and inducing apoptosis

• High levels of Bcl-2 confer resistance to apoptosis

Venetoclax (Venclexta) - Mechanism 

• small molecular competitive inhibitor of Bcl-2

• BH3-mimetic

  • BH3 is a protein domain that mediates the dimerization between anti- and pro-apoptotic proteins

• Releases pro-apoptotic proteins (ex: Bax and Bak) from Bcl-2

• Allows pro-apoptotic proteins to dimerize and induce apoptosis

Venetoclax (Venclexta) - Clinical Use

• chronic lymphocytic leukemia (CLL)

  • CLL patients have a high level of Bcl-2

• AML (acute myeloid leukemia) in combination with chemotherapy

  • Bcl-2 is linked to chemotherapy resistance in AML

Venetoclax (Venclexta) - Side Effects

• tumor lysis syndrome (TLS)

  • caused by the fast breakdown of cancer cells

  • can lead to acute electrolyte and metabolic imbalances

  • potential kidney failure (requires gradual dosing and close monitoring)

  • Patients with impaired kidney function are more susceptible

• Low white blood cell count (Neutropenia)

  • due to inhibiting Bcl-2 in neutrophil precursors

    • neutrophile precursors are very sensitive to Bcl-2 level

  • increased risk of infection

PARP inhibitor - rationale

• poly ADP-ribose polymerase (PARPs) are involved in detecting and signaling cellular responses to single-strand DNA break (SSB) 

• Cancer cells with other DNA repair defects are more reliant on PARP1 to maintain DNA integrity and cell viability 

Olaparib (Lynparza) - Target

PARP1 

• Poly ADP-ribose polymerase 1 

• PARP1 facilitates DNA repair (base excision repair, BER)

Olaparib (Lynparza) - Mechanism

• small molecular inhibitor of PARP1

• Failure of BER causes an accumulation of SSB (single-strand break) and traps PARP1 on DNA

• Promotes DSB (double-strand break) during DNA replication

• Lethal to cancer cells with defects in DSB repair (ex: BRCA1/2 mutations) - unrepaired DSB is lethal to cells

Synthetic Lethality

• condition 1: PARP1 inhibition (PARP1 “defects”) - promotes DSB

• Condition 2: DSB repair defects - due to existing mutations (BRACA1/2)

  • results → Accumulation of too many DBS that become lethal to cells

Olaparib (Lynparza) - Clinical Use 

• ovarian and breast cancers with BRCA mutations 

  • BRCA genes facilitate DSB repair

Olaparib (Lynparza) - Side effect

• Myelosuppression (suppression of bone marrow where blood cells are produced)

  • hematopoietic progenitors (blood cell precursors) in bone marrow are more sensitive to DSB

  • Low white blood cell counts (increased risk of infection)

  • Low red blood cell counts (anemia)

  • patients with underlying bone marrow defects are more susceptible

Olaparib (Lynparza) -  Resistance

• PARP1 mutations that diminish inhibitor binding

• Restoration of other defective DNA repair mechanisms

  • reversion mutations on BRCA (additional mutations in BRCA that restore function)

  • Amplification of wide-type or hypomorphic (partial loss of normal gene function) BRCA

  • Becoming less reliant on PARP1 for DNA repair

Development of Targeted Therapy

• identification of specific molecular targets and vulnerabilities in the cancer cells

• Development of effective therapeutic agents against the candidate targets

• biomarkers for selecting patient populations

• toxicity limits treatment doses and duration

• diverse therapeutic resistant mechanisms

Enabling the acquisition of various cancer hallmarks

• avoiding immune destruction

• tumor-promoting inflammation

Tumor-Promoting Inflammation

• Inflammation is a normal biological response to injury or infection

• Chronic and deregulated inflammation can promote cancer

Inflammation is a normal biological response to injury or infection 

Defenses against infections and promotes repair

Mediated by: 

• immune cells 

  • Neutrophils, macrophages, and lymphocytes (T-cells, B-cells, Natural Killer cells)

• Chemicals released by immune cells, stromal cells, and/or cancer cells

  • cytokines, chemokines, interleukins

  • regulate immune responses (ex: recruit immune cells, activate immune cells)

Chronic and deregulated inflammation can promote cancer

Cytokines, chemokines, interleukins can support various cancer hallmarks

Simplified Hematopoietic Lineage

Inflammatory microenvironment - Cancer promoting effects

Mediated by chemokines, cytokines, and interleukins 

• Angiogenesis 

• Proliferation 

• Metastasis

• Therapy resistance (promote survival)

Inflammatory Microenvironment - Immune cell types

• tumor-associated macrophages (TAMs)

  • secrete pro-inflammatory cytokines

  • promote growth and angiogenesis

• Nutrophils

  • release reactive oxygen species and proteases

  • Promotes invasion and metastasis

Avoiding immune destruction

Immune cells can recognize and destroy/control cancer cells

How do immune cells recognize cancer cells?

• mutant antigens: somatic mutant proteins can be processed and presented as neoantigens

• “Missing-self”: insufficient levels of MHC-I (major histocompatibility complex class I) trigger the “missing-self” signal

• Bound Antibody: Cancer cells bound by antibodies can be recognized

How do cancer cells evade immune surveillance

• suppress antigen presentation

• Suppress immune cell activation

• immunosuppressive tumor microenvironment

Cancer immunotherapy 

harness the immune system capability to recognize and target cancer 

Major immune cell types that directly target cancer cells 

• cytotoxic (CD8+) T lymphocytes (T cells) 

• Natural killer cells (NK cells) 

Cytotoxic (CD8+) T lymphocytes (T cells)

• adaptive immune system

• recognize neoantigens on cancer cells through T cell receptor (TCR)

• Directly kill cancer cells 

Natural killer cells (NK cells)

• innate immune system

• recognize cancer cells through “missing-self” signal and bound antibodies 

• directly kill cancer cells 

Cancer immunotherapy 

1. Immunomodulator

2. Immune checkpoint modulator 

3. Adoptive Immunotherapy 

4. Emerging areas in Immunotherapy 

Example of an Immunomodulator

Aldesleukin 

Examples of a Immune Checkpoint Modulator 

• Ililimumab 

• Pembrolizumab 

Examples of Adoptive Immunotherapy

Tisagenlecleucel (Tisa-cel)

Aldesleukin (Proleukin)

Recombinant human interleukin 2 (IL2) analog

• modified human IL2

Aldesleukin (Proleukin) - Mechanism

• IL-2 is an interleukin that stimulates the growth and differentiation of T cells

• Stimulates the immune system

Aldesleukin (Proleukin) - Clinical use

metastatic melanoma and metastatic renal cell carcinoma

Aldesleukin (Proleukin) - Side Effects 

Allergic reactions 

• rash, itching, swelling, severe dizziness, and trouble breathing

Immune check point modulators

a therapeutic strategy based on modulating the immune checkpoint to promote immune responses against cancer cells

Immune checkpoint

• limits the strength and duration of immune responses

• prevents immune system hyperactivation

• exploited by cancer cells to suppress immune cell activation

T-cell Response to Cancer Cells 

• T cells are major mediators of the immune response against cancer cells 

• T-cell activation:

1. antigen-presenting cells (APCs) (ex: dendritic cells) capture tumor antigens. → presentation 

2. APCs present antigens to and activate naive T cells in the lymphatic tissues → Priming 

3. Cytotoxic T cells recognize antigens on cancer calls and exert cytotoxic effects → Effector 

T-cell Activation 

activation of Naive T cells promotes maturation (Priming) 

activation of mature cytotoxic T cells triggers clonal expansion and cytotoxic effect (Effector) 

Mediated by stimulatory and inhibitor signals

• stimulatory receptor signals

  • T cell receptors (TCR): how T cells recognize antigens

  • CD28

• Inhibitory receptor signals (Immune checkpoints)

  • CTLA-4

  • PD-1

• Activation promotes survival, clonal expansion, differentiation/maturation (naive T cells), and cytotoxic function (cytotoxic T cells)

Immune Checkpoint Signals - Negatively Regulate Immune Responses

• CTLA-4 (Cytotoxic T Lymphocyte-Associated Protein 4)

• PD-1 (Programmed Cell Death Protein 1)

CTLA-4 (Cytotoxic T Lymphocyte-Associated Protein 4)

• Transmembrane glycoproteins are primarily on naive T cells

• Binding to ligand on antigen presenting cells (APC) negatively regulates naive T cell activation

• Compete with CD28 (a stimulatory receptor) for the same ligands (B7)

PD-1 (Programmed cell Death Protein 1)

• Transmembrane glycoproteins are primarily found on mature T cells

• Binding to ligands negatively regulates T cell activation

• The ligand PD-1 or PD-L2 is expressed on some tumor cells, antigen-presenting cells (APC), and other immune cells (ex: tumor-associated macrophages)

Negatively regulate T-cell activation

• checkpoint signal

  • CTLA-4

  • PD-1

• T-cell Activation Phase

  • Priming

  • Effector

• Tissue Location

  • Lymphoid Tissue 

  • Tumor Microenvironment 

Immune Checkpoint Inhibitors

• Block the immune checkpoint signals

• Promote T cell activation

Ipilimumab

A monoclonal antibody against CTLA-4

Ipilimumab - Mechanism

• blocks the CTLA-4 inhibitory signal

  • binds to CTLA-4 and blocks the binding of B7 (ligand)

• Indirectly promotes the CD28 stimulatory signal

  • Available B7 can bind to CD28

• Promote naive T-cell activation at the priming phase in lymphoid tissues

Pembrolizumab 

A monocolonal antibody against PD-1

Pembrolizumab - Mechanism

• Blocks the inhibitory PD-1 signaling

  • binds to PD-1 and prevents the binding of PD-L1 or PD-L2 (ligands)

  • PD-L1 and PD-L2 are expressed on some cancer cells and other cells in the tumor microenvironment

• Promotes T-cell activation primarily at the effector phase in the tumor microenvironment (TME)

Pembrolizumab - Clinical Use

• unresectable and metastatic melanoma (Iplilimumab and Pembrolizumab)

• Other advanced stage cancers (Pembrolizumab)

• Micro satellite instability-high (MSI-H) or mismatch repair deficient (dMMR) cancers (High mutation loads)

Pembrolizumab  - Side Effects

• Autoimmune-like symptoms

• Damages in normal tissues and organs

Immune Checkpoint Inhibitors 

Other Immune checkpoints as potential therapeutic targets 

• LAG-3 (Lymphocyte activation Gene-3)

• TIM-3 (T cell immunoglobulin and mucin-domain caontaining-3)

• TIGIT (T cell immunoglobulin and ITIM domain)

• VISTA (V-domain Ig suppressor of T cell activation)

• BTLA (B and T lymphocyte Attenuator) 

Adoptive Immunotherapy

Chimeric Antigen Receptor T Cell (CAR-T cells)

Tisagenlecleucel

Tisagenlecleucel

• Engineer T cells from patients to recognize and attack B-cell malignancies in the same patient 

• Some B cell malignancies have increased levels of CD19

Tisagenlecleucel - Procedures

• Isolate T cells from patients

• Genetically engineer the T cells to express a Chimeric Antigen Receptor (CAR) that recognizes CD19

• Expand and enrich these engineered T-cell population

• Infuse into the same patients (autologous)

Tisa-cel - clinical use

• relapsed B-cell acute lymphocytic leukemia (ALL)

• relapsed diffuse large B-cell lymphoma (DLBCL)

Tisa-cel - Side effects

• Cytokine-release syndrome (CRS)

  • massive release of cytokines due to T cell activation

  • Rapid heartbeat, low blood pressure, and trouble breathing

• Risk of developing T-cell malignancies

Limitations of CAR-T Cell Therapy 

• T-cell exhaustion 

• Heterogenous tumor populations 

• Challenges in solid tumors 

• complex personalized procedures 

• Severe cytokine-released syndrome (CRS)

Advances in Chimeric Antigen Receptor Design

• Co-stimulatory signaling domains in newer generations of CAR

• Improve T cells expansion and survival 

New CAR designs

• Bispecific CAR

  • bind multiple antigens

  • Diminish antigen escape

• Tunable and regulate CAR

  • Kill switch in the event of severe toxicity

  • Reversible switch to prevent T cell exhaustion

• T Cell Receptor (TCR)-like CAR

  • CAR: Surface protein, MHC-independent

  • TCR (native T cells): Processed intracellular proteins, MHC-dependent

Emerging areas in Immunotherapy

• Tumor-infiltrating lymphocyte

  • Lifileucel

• Immune cell engager

  • Blinatumomab

• Natura killer (NK) Cells

Tumor Infiltrating Lymphocytes

Lifileucel

Lifileucel - Procedure

• isolate tumor-infiltrating lymphocytes (TILs) from a patient’s tumor

• Expand the TIL populations in facilities

• Infuse back to the same patient (autologous)

• No cell engineering (distinct from CAR-T)

• First cellular therapy to be approved for a solid tumor (Melanoma)

Blinatumomab

Bispecific T-cell engager

Blinatumomab - Mechanism

• Bispecific antibody engaging T cells to cancer cells 

• CD19-directed CD3 T-cell engager

  • CD3 on T cells

  • CD19 on malignant B cells

Blinatumomab - clinical use

CD19-positive B-cell acute lymphoblastic leukemia (ALL)

Blinatumomab - side effects

Cytokine Release Syndrone (CRS)

natural Killer (NK) cells

• Recognize cancer cells by:

  • “missing-self” signals 

    • imbalance between stimulatory and inhibitory signals 

    • insufficient levels of MHC-I on cancer cells leads to lowered inhibitory signals and activation of NK cells 

  • Antibody-dependent cell-mediated cytotoxicity (ADCC)

    • Bound antibodies on cancer cells are recognized by NK cells through a surface receptor CD16

Harnessing NK Cells to Target Cancers - Immune cell engagers 

• Bispecific antibody (NK cell engagers)

  • CD19 on NK cells

  • Antigens on cancer cells

• CAR-NK

  • Chimeric Antigen Receptor (CAR)

    • similar concepts to CAR-T but distinct design

Harnessing NK Cells to Target Cancers - Advantages 

• allogenic 

  • do not require cells from the same patient

  • able to utilize NK cells from donors 

• Cytokine release syndrome (CRS) is less common 

  • NK cells have a lower cytokine response than T cells 

Harnessing NK Cells to Target Cancers - Limitations 

• Shorter lifespan (1-2 weeks) 

  • CAR-T can last for months to years 

• Tumor infiltration and suppressive tumor environment (similar to CAR-T)

Personalized (precision) Medicine

• a way health care providers can offer and plan specific care for their patients, based on genes, proteins, and other substances in a person’s body

  • With regards to cancer, precision medicine most often means looking at how changes in certain genes or proteins in a person’s cancer cells might affect their care, such as their treatment options

Personalized (precision) medicine - ultimate goal

to shift healthcare from a one-size-fits-all approach to a more individualized, precise, and effective method of diagnosis, treatment, and prevention, improving patient outcomes and reducing adverse effects

Diagnostic techniques in personalized medicine

Major techniques in cancer diagnosis

1. DNA/RNA sequencing

2. Proteomics

3. Immuno-profiling

4. Imaging

DNA sequencing in cancer detection - Tumor profiling 

Involves tumor biopsy for either targeted or whole genome sequencing 

• categorize mutations enriched in specific cancer types and potentially primary tumor sites vs. metastatic sites 

Pharmacogenomics in cancer therapy

Pharmacogenomics is important to understand an individual’s genes affect how they respond to specific medications, which may vary cancer treatment outcomes

Development of patient-derived models for preclinical studies

Patient-derived cancer methods

• Patient tumor biopsy can be used to generate ex vivo cancer models

  • cell lines - 2D in vitro culture

  • Tumor organoids - 3D in vitro culture

  • Tumor xenografts - typically in an immunocompromised mouse

• used for genome sequencing to biomarker analyses

• Patient-specific cells to determine new drug or drug combination for treatment options

Minimal invasive method for cancer detection and therapeutic monitoring - liquid biopsy 

Typically, blood samples

• minimally invasive 

• can determine appropriate treatment 

• monitor cancer development 

• track response to treatment 

• assess rick for cancer progression/resistance 

Imaging techniques in cancer detection - Antibody-radioisotope labeling

used to detect biomarker-positive cancer sites

What is the main goal of clinical trials?

Determine the safety and efficacy of the investigational new drug (IND)

Phase 1 - Clinical Trial

Safety

• is the investigational medication/treatment safe?

  • are there side effects 

  • how does it affect or move through the body

  • is it safe to use at the same time as other medications

• Who is in it

  • small group of healthy people, generally less than 100

Phase 2 - Clinical Trial

Efficacy

• Is the investigational medication/treatment effective in treating the targeted condition?

  • Does the treatment relieve, reverse or stop the progression of the condition?

  • How safe is it?

  • What is the most effective dosage?

• Who’s in it?

  • generally 100-300 people with the exact condition being studied

Phase 3 - Clinical Trial

Confirmation

• How does the investigational medication/treatment compare to the standard treatment for the condition?

  • more effective, less effective, the same?

  • Longer-term adverse effects?

  • How does it affect quality of life, or survival?

  • How might it be used along with existing treatments?

• Who’s in it?

  • Often 300-3,000 people with the exact condition being studied

Phase 4 - Clinical Trial (post-FDA approval)

Follow Up

• After the investigational medication/treatment is approved, how does it work for other patients with the condition?

  • More safety/efficacy information is gathered

  • are there long-term benefits?

  • are there long-term risks?

• Who’s in it?

  • often several thousand people who have been prescribed the investigational medication

Basket Trials

prospective clinical trials that test one or more targeted interventions across multiple types of diseases 

Umbrella Trials

Prospective clinical trials that test multiple targeted interventions for a single disease based on predictive biomarkers or other predictive patient risk factors