An article combs: EGFR/ALK TKI's past and present lives and the progress of third-generation drug research

The discovery of EGFR mutations and ALK rearrangement drives biomarker-oriented treatment patterns
in advanced NSCLC.
Several EGFR and ALK tyrosine kinase inhibitors (TKIs) have been used in the clinic, and the latest drugs in the clinic are third-generation EGFR TKI and ALK TKI, but drug resistance remains an unsolved fundamental challenge
.
There are two main mechanisms of resistance: "on-target" (mainly mediated by acquired drug-resistant mutations in the EGFR or ALK kinase domains) and "off-target" (mediated by non-target kinase mutations, such as bypass signaling pathway activation or phenotypic transformation).

Not long ago, a review in the journal NATURE REVIEWS CLINICAL ONCOLOGY detailed the development history of third-generation drugs, the mechanism of drug resistance and the development direction
of new drugs.

EGFR mutation in NSCLC

EGFR is a member of the family of
receptor tyrosine kinase (RTK) ErbB.
Binding of ligands to extracellular domains of EGFR can cause conformational changes that lead to receptor dimerization, tyrosine phosphorylation, and activation of downstream signaling pathways (RAS–MAPK, PI3K–AKT, and JAK–STAT), which in turn affects cell growth and survival
.
Although EGFR is also expressed in a range of non-malignant tissues, it is overexpressed (at least 5-fold) in multiple epithelial cancers and has long been considered a potential therapeutic target
.

Development of EGFR TKI – the first and second generation of EGFR TKI

First-generation EGFR TKI has shown only general antitumor activity in patients with unselected advanced NSCLC, with an objective remission rate (ORR) of 10 to 20% and a median progression-free survival (PFS) of 3 months
.
But higher ORR
was observed in patients who did not smoke and Asia.
Subsequent translational studies have found that somatic carcinogenic changes in the EGFR kinase domain are enriched
in both groups of people.
The most common types of activated EGFR mutations are exon 19 deletion and L858R point mutations, which are 100-fold more sensitive to EGFR TKI than wild-type
receptors.
Prospective trials followed up quickly and found that gefitinib or erlotinib was at an unprecedented level in this group of people, with an ORR of approximately 75% and a median PFS of 7 to 12 months
.

However, first-generation EGFR TKI typically develops acquired resistance within a year, and the most common mechanism of resistance is T790M, which occurs in about 60% of patients
.
To overcome the EGFR T790M resistance mutation, the second and third generations of EGFR TKI came into being
.
Compared to the first generation of EGFR TKI, the second generation of EGFR TKI improves the median overall survival (OS) of patients by about 8 months
.

Third generation EGFR TKI

Third-generation EGFR TKI ositinib is more
active and selective against EGFR-sensitive mutations and T790M drug-resistant mutations.
Oshitinib's decreased affinity with certain drug transporters results in better retention in the cerebrospinal fluid after crossing the blood-brain barrier, so ositinib shows excellent ORR and PFS in patients including the central nervous system (CNS
).
In 2015, osirtinib was accelerated by the FDA for use in patients with EGFR T790M-positive NSCLC who have progressed treatment with
previous EGFR TKI.
Subsequently, based on the results of the FLAURA study, ositinib was also approved by the FDA for first-line treatment
in patients with advanced NSCLC with EGFR-sensitive mutations.
Compared with the first generation of EGFR TKI, ositinib can significantly prolong the median PFS and median OS in patients, and the incidence of CNS progression is reduced
.
Other third-generation EGFR TKIs such as amettinib and vometinib have also shown good antitumor activity and safety, and are being further followed up and explored
.
Partially approved EGFR TKI

ALK rearrangement in NSCLC

ALK rearrangement was first discovered in NSCLC in 2007, and since the EML4-ALK rearrangement was first discovered, more than 90 ALK fusion mates
have been identified in NSCLC.
Mechanically, ALK rearrangement leads to the constitutive activation of ALK kinase and related downstream cell signaling pathways such as RAS-MAPK, PI3K-AKT, and JAK-STAT, leading to proliferation and survival
of tumor cells.

First and second generation ALK TKI

Clinically, ALK rearrangement is associated
with adenocarcinoma, younger diagnosis, and less smoking.
Crizotinib was the first targeted drug to be evaluated in ALK rearrangements patients, originally developed as a broad-spectrum TKI of MET inhibitors, but found to be a potent ALK/ROS1 inhibitor
.
Phase I/II studies have shown that crizotinib can have an ORR of up to 60%
in ALK rearrangement NSCLC.
In 2013, crizotinib was approved by the FDA for ALK rearrangement of advanced NSCLC, marking a major paradigm shift
in biomarker-guided therapies that can be extended beyond EGFR mutations and support the individualized management of advanced NSCLC.

As with EGFR TKI, crizotinib develops acquired resistance, usually within one year of initiation of treatment (median PFS of 7.
7 to 10.
9 months
).
Subsequently, researchers developed a more powerful second-generation ALK TKI, including cerratinib, alectinib, and bulgertinib
.
Second-generation ALK TKI has good activity in crizotinib-resistant patients and improves CNS activity
.
Partially approved ALK TKI

Third generation ALK TKI

Loratinib is a third-generation, potent and highly selective ALK/ROS1 TKI developed for early ALK TKI-acquired drug resistance with broad-spectrum anti-ALK kinase resistance mutant activity
.
Based on related studies, loratinib was approved by the FDA in 2018 for the use of first-line crizotinib or cerratinib or crizotinib sequentially through the progress of other ALK TKI treatments for ALK rearrangement of advanced NSCLC
.
Based on the phase III CROWN study, first-line lolatinib was significantly superior to crizotinib, with a 72% reduction in disease progression or mortality, and loratinib also significantly prolonged CNS disease progression, based on which in 2021 loratinib was approved by the FDA for ALK rearrangement in patients with advanced NSCLC, including first-treatment patients
.

Mechanisms of acquired drug resistance

Mechanisms of acquired resistance to third-generation EGFR and ALK TKI

The "on-garget" resistance mechanism of ositinib

Oshitinib binds to EGFR by forming a covalent bond with cysteine 797 (C797) residues located in the ATP binding fissure
.
Thus, the C797 residue is a vulnerable site
for the development of acquired resistance to ositinib.
In fact, the EGFR C797S mutation is seen in 10% to 26% of patients with
second-line treatment progression to ositinib.
Other gene mutations that can lead to ositinib resistance have a lower incidence than C797S, including G796X, solvent frontier mutations L792X, L718X, G724S
.


Nevertheless, the cumulative incidence of these mutations is much lower than the T790M mutation (50%-60%)
that causes ositinib resistance.
Compared with second-line oxitinib treatment, oxitinib "on-target" resistance appears to be less common
in first-line therapy.
In the FLAURA study, only 7% of patients developed the EGFR C797X mutation, and as expected, the EGFR T790M mutation
was not detected in ositinib-resistant tumors.

"On-garget" resistance mechanism of loratinib

ALK kinase domain mutations lead to acquired drug resistance to second-generation ALK (approximately 50%-60% of patients), and each second-generation ALK TKI is associated
with a unique ALK resistance mutation.
For example, the most common ALK-resistant mutations for alitinib are G1202R (25%–30% of patients) and I1171X (10%–15%)
.
In fact, the main causes of acquired resistance to loratinib are compound ALK mutations (e.
g.
, C1156Y/L1198F, G1202R/L1196M, I1171N/D1203N, etc.
).

Whole exome sequencing and analysis suggest that drug-resistant mutations can accumulate gradually, eventually leading to highly refractory compound mutations that are more therapeutically challenging
after continuous treatment with ALK inhibitors.

"off-target" resistance mechanisms

As third-generation inhibitors become more widely available in first-line treatments, the occurrence of "off-target" resistance is expected to increase
.
The "off-target" resistance to EGFR/ALK TKI, although substantial progress has been made in theory, is still unknown in a considerable area (40% to 50% of patients after first-line ositinib) and further research
is needed.

Parallel bypass activates the signal path

A key point of drug resistance is attributed to the activation of the bypass signaling pathway, which must be accompanied by both inhibition of driver gene mutations and bypass activation signaling pathways
in order to successfully overcome drug resistance.
MET amplification is the first resistance mechanism to be found in the EGFR mutation NSCLC involving a bypass activation signaling pathway and is seen in 7%-15% of patients receiving ositinib first-line and 9.
8%-30% of patients receiving
ositinib at the post-line line.
Importantly, combination therapies using targeted MET inhibitors are clinically feasible
.
For example, the TATTON study evaluated the efficacy of ositinib + MET TKI cervotinib, and the results were encouraging, with an ORR of 30%
in patients with MET amplification and EGFR mutations.
Other mechanisms that have been identified for bypass-activated resistance are shown in the table below
.


Table EGFR mutation NSCLC bypass signaling pathway

Downstream signaling pathways

Reactivation of the EGFR and/or ALK downstream signaling pathway can lead to drug resistance
.
For example, the RAS-MAPK signaling pathway is a key downstream pathway for EGFR and ALK and can be reactivated by signals from multiple nodes (e.
g.
, acquired BRAF fusion, KRAS mutation, NRAS mutation, MAP2K1 mutation, DUSP6 deletion, wild-type NRAS or KRAS increase).

Preclinical studies suggest that regulation of the ERK signaling pathway can lead to resistance to third-generation EGFR TKI, combined inhibition of MEK/ERK or recovery of sensitivity
to EGFR TKI.

Histological transformation

Resistance to early-stage EGFR inhibitors can lead to transformed small cell lung cancer (SCLC), which accounts for 3% to 14%.

First-line ositinib may be more common
after treatment.
EGFR mutations persist in transformed tumor samples but no longer rely on the EGFR signaling pathway, resulting in ineffective EGFR TKI
.
One retrospective study found that patients with EGFR mutant SCLC had worse outcomes compared to patients with EGFR mutation NSCLC, including response to chemotherapy and OS outcomes
.
Studies have also found that after first-line ositinib treatment, patients can also be converted from adenocarcinoma to squamous cell carcinoma
.
Translational small cell lung cancer and squamous cell carcinoma may also occur after the progression of ALK TKI therapy
.
Re-biopsy
after disease progression is highly recommended.

Epithelial interstitial transformation (EMT)

EMT is detected in both EGFR mutations and ALK rearrangement patients and is thought to promote tumor invasion
.
Over the past few years, EMT has been thought to be associated
with oshitinib and loratinib resistance.
In the case of EMT-mediated ositinib resistance, the ATR-CHK1-aurora B signaling cascade is activated and EGFR inhibitors in combination with auroraB kinase may overcome the resistance
.

Management of progression of systemic diseases

For patients with diffuse, systemic disease progression, treatment options can be broadly divided into alternative TKI (designed to overcome "on-target" resistance) and strategies other than TKI (aimed at overcoming "off-target" resistance
).

The new generation of TKI

Several fourth-generation EGFR TKIs and ALK TKIs are currently under development with the goal of overcoming the "on-target" resistance of third-generation TKIs
.
EAI045 is an allosteric EGFR inhibitor, and preclinical studies have shown that EAI045 combined with cetuximab can successfully inhibit the growth
of L858R-T790M mutation and T790M-T790M-C797S mutant lung cancer.


JBJ-04-125-02 is another allosteric EGFR inhibitor currently under development and has been found to be more effective when JBJ-04-125-02 inhibits L858R–T790M–C797S mutations, JBJ-04-125-02 may be more effective
when combined with oshitinib.

Recently, the fourth-generation EGFR TKI BLU-945 and BLU-701 have shown strong CNS penetration
.
In preclinical studies, BLU-945 has been shown to inhibit the trimutant EGFR subtype (EGFR L858R mutation, exon 19 deletion, T790M mutation, and C797S mutation), while BLU-701 is active
against dual mutants (EGFR-sensitive mutation and C797S).
BLU-945 and BLU-701 are currently in early clinical trials
.

The new ALK TKI is also currently under
development.
TPX-0131 is a new generation of ALK TKI with a compact large ring structure that can fully bind to the adenine site in the ATP pocket, inhibiting the ALK signaling pathway
even in the presence of ALK-resistant mutations.
NVL-655 is a selective, CNS-penetrating next-generation ALK TKI that is effective in the G1202R-L1196M, G1202R–L1198F, and G1202R–G1269A complex mutations and is currently under clinical study
.

Recycling of TKI

In addition to the development of novel ALK TKIs, the "reuse" of older generation TKI or broad-spectrum kinase inhibitors with potential effects may overcome the "on-target" resistance of third-generation EGFR/ALK TKI
.
For example, bulgetinib + cetuximab was found to inhibit three mutations of EGFR and to inhibit ositinib resistance
.


Certain loratinib-resistant mutations ALK rearrangement NSCLC may be sensitive
to older generations of ALK TKI.
For example, one patient was resistant to crizotinib due to ALK C1156Y mutation, and subsequently resistant to lolatinib due to the appearance of ALK C1156Y–L1198F, and studies have shown that the ALK C1156Y–L1198F mutation is sensitive to crizotinib, enabling patients to successfully reuse crizotinib
.

Combination therapy

Combination therapy is an alternative treatment strategy that overcomes or delays resistance to third-generation inhibitors
.


One resistance strategy includes the early generation and the third generation of EGFR TKI, which, in combination, overcomes or delays the occurrence
of "on-target" resistance.
For example, in the EGFR mutation NSCLC, ositinib in combination with first-generation EGFR TKI overcomes the resistance-associated C797S mutation
.
Niederst et al.
demonstrated that when the EGFR T790M mutation and the C797S mutation appear trans (located in different EGFR alleles), the first generation of EGFR TKI (targeting C797S) and the third generation of EGFR TKI (targeting T790M) can restore the inhibition of the EGFR signaling pathway
.

A Phase II ORCHARD study is currently evaluating the efficacy
of ositinib + gefitinib in patients with EGFR C797X mutations who progress to first-line oxitinib therapy.
Two centres in Australia are conducting a study to evaluate the efficacy of alternating crizotinib and loratinib with the aim of delaying the development
of "on-target" resistance.

The second joint strategy is EGFR/ALK TKI in combination with another targeted drug designed to overcome "off-target" resistance
.
Similarly, in patients with ALK rearrangement of NSCLC, multiple studies are underway in combination with other targeted therapies
.

Table 2 Research progress related to third-generation drug combination therapyThe third combination strategy is EGFR/ALK TKI+ chemotherapy
.
The Phase III COMPEL study aims to assess whether platinum plus pemetrexed plus ositinib should be continued concomitantly after first-line ositinib
in the treatment of extracranial disease progression.
Gefitinib + chemotherapy significantly prolongs PFS and OS in patients with EGFR mutation NSCLC compared to gefitinib
.
The Phase III FLAURA2 study is exploring the efficacy
of oxitinib+ chemotherapy versus ositinib for the initial treatment of patients with non-squamous NSCLC.

New therapies and future directions

Antibody-conjugated drugs (ADCs) and bispecific antibodies are novel therapies currently under development that may be effective
in patients with TKI resistance.
Amivantamab is a bispecific antibody targeting EGFR and MET based on the CHRYSALIS study that was accelerated for use in patients with
EGFR exon 20 insertion mutation NSCLC in May 2021.
In addition, amivantamab in combination with third-generation EGFR TKI lazertinib has shown initial efficacy in patients with oximinib progression, with an ORR of 36%.


Patritumab deruxtecan is an HER3-targeting ADC with antitumor activity
in patients with at least one EGFR TKI progression (86% of patients receiving ositinib).
Patritumab deruxtecan has been recognized as a single drug by the FDA as a breakthrough therapy for use in patients with third-generation tyrosine kinase inhibitors and patients
with drug-resistant EGFR mutation metastasis or locally advanced NSCLC during or after platinum-containing chemotherapy.
The efficacy of patritumab deruxtecan monotherapy or in
combination with the treatment of patients with EGFR mutations is being explored.

Other innovative treatment strategies, including cancer vaccines, are also expected to treat patients with
EGFR mutations and ALK rearrangement NSCLC.
An ALK vaccine has shown a strong immune activation response in preclinical studies, laying the foundation
for exploring innovative strategies for anti-ALK vaccines.

The discovery of EGFR-sensitive mutations and ALK rearrangement driver genes has set a precedent for the transformation of treatment mode in advanced NSCLC
.
However, drug resistance remains a challenge that has not been fully addressed
.
Current collaborations in basic, translational, and clinical research will provide a deeper and broader understanding of the mechanisms underlying the biology of TKI-resistant cancers, enabling the rational development of innovative therapies
.

Editor: Xiaoyuan Typesetting: Xiaoyuan Execution: Xiaoyuan

References: Cooper AJ, Sequist LV, Lin JJ.
Third-generation EGFR and ALK inhibitors: mechanisms of resistance and management.
Nat Rev Clin Oncol.
doi: 10.
1038/s41571-022-00639-9.
Epub ahead of print.
PMID: 35534623.