Molecular Profiling for Patients with Solid Tumors: A Single-Institution Experience

Molecular Profiling (MP) of tumors is innovative progress that led to identifying targetable alterations that could be exploited to deliver personalized cancer treatment. Lack of data from the region about the clinical utility of has prompted this study. Tumor tissues from 100 consecutive adult patients with solid tumors were genomically profiled successfully using commercially available platforms. Outcomes for patients who received an MP-guided versus MP-unguided therapy were compared. Progression-Free Survival (PFS) was the primary endpoint, while Overall Survival (OS) was the secondary endpoint. Patients’ median age was 57 years, and female patients constituted 65% of the series. Thirty-one patients were newly diagnosed, and 69 patients had the MP performed upon disease recurrence or progression. Breast, lung, and colorectal cancers were the most frequent tumors. In 90 of the tested tumors, one or more aberrations were identified. In 61 patients, the MP results suggested at least one matched agent and guided therapy in 53 patients. Of all patients who received further therapy (83 patients), the median PFS was significantly longer in patients whose MP-guided versus those whose treatment was not guided (21.8 [95% CI; 14.5 - 29.1] vs. 10.9 [95% CI; 6.2 - 15.6] months, hazard ratio [HR] = 0.34 [95% CI; 0.17 - 0.69], P = 0.002). The benefit was largely shown in patients with recurrent or progressive disease (HR = 0.32 [95% CI; 0.14 - 1.20.75]; P = 0.006). While patients who received MP-guided therapy had numerically higher OS rates, that difference was not significant. This preliminary experience demonstrated MP’s feasibility for cancer patients with a significant improvement in PFS, albeit a lack of OS benefit. Further research is warranted to address the inherent challenges for the universal adoption of MP in daily practice.


Introduction
In the present era of precision medicine, recent advances in molecular cancer biology have led to the identification of tumorspecific molecular aberrations that could be exploited to inform tumor diagnosis, prognosis, and drive therapeutic decisions by precisely targeting such aberrations [1][2][3].
Currently, there is no uniformity concerning the clinical utility of comprehensive Molecular Profiling (MP) in patients with solid tumors, and the only randomized trial conducted so far showed no benefit of MP-guided targeted therapy. In that phase II SHIVA study, Le Tourneau et al. randomized 197 pretreated patients with solid tumors to receive a matched molecularly targeted agent or treatment at a physician's choice [4]. No Progression-Free Survival (PFS) difference was demonstrated; however, in a recent update, it was concluded that patients who crossed over from the control arm to the experimental arm achieved a 30% improvement in PFS [5].
Another study, the My Pathway trial, evaluated the effectiveness of several targeted therapies in 35 tumor types (in 230 patients) that harbor genetic alterations not labeled for such treatments [6]. The study concluded that the approved targeted therapy regimens achieved responses in several refractory solid tumor types that were not labeled for these agents.
We recently reported our preliminary analysis of the MP of 50 consecutive adult patients with solid metastatic cancers refractory to standard of care [7]. The median PFS was improved among those whose therapy decision was guided by the MP findings (12.0 months) compared with those whose MP could not recommend a specific management decision (5.2 months). While there was no significant Overall Survival (OS) difference, the 12 month OS rate was 64% vs. 53%, respectively.
The current study aimed to provide a more mature analysis and a longer follow-up of prospective, comprehensive MP of tumors from a series of 100 consecutive patients with solid tumors using Next-Generation Sequencing (NGS). There has been no data about the use of tumor profiling among cancer patients in Saudi Arabia or the nearby countries to the best of our knowledge. Moreover, there are no local institutional or national guidelines that direct clinicians to exploit the emerging technology to achieve a better patient outcome.

Patients
Between May 2017 and April 2020, the first 100 consecutive adult patients with solid tumors whose tumors were tested for MP were included in the current analysis. Molecular profiling was requested for several patient groups: at diagnosis for those whose tumors are associated with benefit from known targetable agents; e.g., lung cancer, those whose initial diagnosis is linked to a poor prognosis, or patients presented with metastatic disease. MP was also performed for patients who demonstrated recurrence, progression, or refractoriness to the care standard. MP was only performed for patients with an acceptable performance status to permit further therapy (Eastern Cooperative Oncology Group performance status 0 to 2). Molecular Profiling (MP) of tumors is innovative progress that led to identifying targetable alterations that could be exploited to deliver personalized cancer treatment. Lack of data from the region about the clinical utility of has prompted this study. Tumor tissues from 100 consecutive adult patients with solid tumors were genomically profiled successfully using commercially available platforms. Outcomes for patients who received an MP-guided versus MP-unguided therapy were compared. Progression-Free Survival (PFS) was the primary endpoint, while Overall Survival (OS) was the secondary endpoint. Patients' median age was 57 years, and female patients constituted 65% of the series. Thirty-one patients were newly diagnosed, and 69 patients had the MP performed upon disease recurrence or progression. Breast, lung, and colorectal cancers were the most frequent tumors. In 90 of the tested tumors, one or more aberrations were identified. In 61 patients, the MP results suggested at least one matched agent and guided therapy in 53 patients. Of all patients who received further therapy (83 patients), the median PFS was significantly longer in patients whose MP-guided versus those whose treatment was not guided ( MP was performed on archival fixed formalin paraffin-embedded tissue either from the primary tumor site or from a metastatic lesion if feasible using either of two commercially available NGS platforms, i.e., Foundation One (Foundation Medicine, Inc.) or OncoDEEP (OncoDNA, Inc). The NGS mutational analysis using the Foundation One was based on a panel of genetic mutations in 324 genes and two genomic signatures in any solid tumor [8]. In comparison, the OncoDEEP platform is based on NGS of 75 cancer-related genes, besides several immunohistochemistry tests, including protein phosphorylation to study protein expression [2].

Decision-making
The findings of the MP for each patient were discussed at a multidisciplinary molecular tumor board to recommend further management. A treatment was considered "matched" if there was an agent(s) that could target an aberration in a patient's MP or a functionally active protein expressed in the tumor and guide therapy decision. Upon obtaining the MP results, the treatment decision was either: 1) uphold the current treatment, or recommend changing/ initiating a different treatment as guided by the MP results; or 2) uphold the current treatment, or recommend changing/starting another treatment not driven by the MP results. Implementing the first or the second recommendation was considered as either an MPguided or MP-unguided decision, respectively.

Statistical methodology
PFS was defined as the interval between the date of implementing a management decision based on MP to the date of progression or death of any cause, whichever came first. OS was defined as the interval between the date of implementing a management decision based on MP to the date of death of any cause or date of the last follow-up. Survival functions were estimated using the Kaplan-Meier method, and survival between groups was compared using the logrank test. All tests were two-sided at the 5% significance level. All statistical analyses were done with SPSS statistical package (IBM SPSS Statistics for Windows, version 25.0., New York, USA).

Results
The median age (95% CI) in years for the entire population was 57 (54 -60) years, with males on average older (60 [53 -67]) than females (56 [52 -60]). Table 1 shows patients and disease characteristics. More females than males (65% v 35%), breast cancer, lung cancer, colorectal cancer, and pancreatic cancer were the most common primary tumor sites. In 31 patients, MP was performed at initial diagnosis, while in 69 heavily pretreated patients, it was done upon disease recurrence or progression.
The median interval (95% CI) between initial diagnosis and the MP was 10.2 (6.1 -17.7) months. Table 2 depicts the summary results of the performed molecular testing. Almost a quarter of the tumors showed PD-L1 expression of >1%, and an Immunohistochemistry (IHC) positivity for CD 8, while microsatellite instability and tumor mutational burden of >10 megabases were uncommon, 2% and 3%, respectively. In 90% of the tested tumors, one or more aberrations were identified by NGS, while in 50 tumors were tested by IHC for protein expression, 41 showed one or more aberrations. Table 3 shows most of the identified aberrations with TP53, PIK3CA, and RAS mutations being the most frequently detected. Table 4 depicts the entire population's management decision, newly diagnosed patients, and those with recurrent or progressive disease. In 61 patients, the MP results suggested at least one matched agent. MP guided therapy decision in 53 of those 61 patients (87%),  Six of the newly diagnosed patients received no further therapy; of those, two patients with metastatic pancreatic cancer, and two with metastatic lung cancer were not fit for or declined systemic chemotherapy. None of those patients had potential matched therapy. Two patients with advanced ovarian cancer were not candidates for Poly (ADP-ribose) polymerase inhibitors maintenance as they had homologous recombination proficient with or without BRCA wild tumors. Of the 69 patients with recurrent or progressive disease, 11 patients were not given further therapy due to deterioration in performance status (8 patients) or loss to follow-up (3 patients). Eight of these 11 patients had potential matched treatment.

Progression-free survival
The database was looked on April 30, 2020, and the median follow-up from the date of MP testing was 16.0 (95% CI; 12.7 -19.4) months. Five, 61, and 34 patients were alive with no evidence of disease, alive with disease, and dead, respectively.
All PFS dates were based on the computation of PFS from the date of MP testing. After excluding patients where no further therapy was offered, the median PFS was significantly longer in patients whose treatment with guided by MP versus those whose treatment was not guided On the other hand, among patients with recurrent or progressive disease, there was a significant difference in median PFS between patients treated as guided by MP results as compared with that among patients whose treatment was not guided ( After excluding patients where no other therapy was given, there was a trend of an improved median OS among patients whose received MP-guided therapy versus those whose treatment was not guided Among the newly diagnosed patients, the median OS has not been reached; however, the 12-and 24-month OS rate (± standard error) was 76% (8%) and 70% (9%), respectively. In this group, after excluding those who were not given any further treatment, there was no difference in the median OS between those who received MPguided versus MP-unguided therapy ( Table 5 showed the OS probability rates of patients according to their status and if the MP results guided a management decision. Numerically, the OS rates for patients who received their treatment as MP-guided achieved higher survival rates as compared with those whose therapy was unguided. That advantage was shown in newly diagnosed patients and in patients with disease recurrence or progression.

Discussion
In 90% of our patients, at least one aberration was identified. The prevalence of detected aberration is influenced by the population examined, the method used, and the number of aberrations intended to be examined. Tsimberidou et al. using a polymerase chain reaction, detected at least one aberration in 40% of patients [9]; on the other hand, using the NGS method, Wheler et al. tested 339 patients and detected a potentially actionable target in 94% of patients [10].
In the current series, tumors in 61 patients identified potential matched agent(s), and in 53 patients (87%), the identified therapy was used. The rate of implementing matched therapy in our series was relatively higher than those reported from other studies [10,11].
Applying the MP guidance achieved a PFS advantage with a 66% reduction in the risk of progression or death (HR = 0.34). The demonstrated benefit was almost identical to that shown in our earlier series [7]. The advantage was evident, particularly in patients with recurrent or progressive disease. On the other hand, no significant OS difference was shown between the MP-guided and the unguided groups.
Nevertheless, as shown in Table 5, the OS rates were numerically higher among those whose treatment decision was MP-guided. The increased OS rates were observed in newly diagnosed patients and among those with disease recurrence or progression. Some several plausible reasons and limitations may explain the lack of OS advantage. First, we only analyzed a small sample of 100 patients, which may have precluded the demonstration of OS advantage. Second, this series included a diverse patient population of different tumor types, besides the inclusion of newly diagnosed patients and patients with disease recurrence or progression. Third, in 39 patients, no targetable aberration was identified. Lastly, intervention postprogression was not controlled and was implemented according to the individual physician's choice. Despite those limitations, our series represents the only available data concerning prospective MP from Saudi Arabia and perhaps from the entire Middle East region to the best of our knowledge.
It is prudent to acknowledge that larger series have also reported conflicting evidence. In a nonrandomized phase I trial, conducted by the MD Anderson Cancer Center and included 1,144 patients with advanced cancer, it was found that patients with one aberration who

Management decision
No.

Guided by molecular profiling 53
Uphold current therapy 24

Change/initiate therapy 29
Not-guided by molecular profiling 30

Uphold current therapy 23
Change/initiate therapy 7

No further therapy 17
Management decision according to the timing of molecular profiling testing

New diagnosis 31
Guided by molecular profiling 17 Not guided by molecular profiling 8

No treatment 6
Recurrent or progressive disease 69

Guided by molecular profiling 36
Not guided by molecular profiling 22  received matched therapy demonstrated higher objective response and superior survival compared with that among those treated with non-matched drugs. However, the advantage was not shown when 2 or 3 molecular alterations were present [11].

No treatment 11
The PREDICT-trial enrolled 347 patients with advanced solid tumors, and a quarter of patients were treated according to their genomic profile [12]. Improvement in PFS was demonstrated in patients treated with a matched therapy compared with the control group; however, no improvement in OS was observed.
One of the most compelling evidence was reported by Schwaederle et al. [13]. The authors conducted a meta-analysis of 570 phase II single-agent studies, mainly nonrandomized (> 80% of the studies), and incorporating 32,149 patients. The meta-analysis concluded that using a personalized strategy was associated with a higher response rate and longer PFS and OS as compared with the non-personalized approach.
Despite the quantum leap progress in our understanding of tumor biology and the resulting evolution of MP, several challenges face the wider use of MP in the daily practice. While MP's results may identify a targetable aberration, the known tumor heterogeneity could certainly influence the clinical outcome [14]. Additionally, there are several other challenges: the uncertainty surrounding whether the complex MP report influences the oncologist's treatment decisions, lack of confidence of physicians in genomic knowledge and how they could interpret MP reports, uncertainty related to the clinical utility of the information, and undoubtedly the associated economic burden [15][16][17].

Conclusion
Although there have been clear successes in the era of molecular characterization, MP's clinical utility remains unproven. Precision medicine among cancer patients remains a major challenge for the oncology community but could enhance more therapeutic options to be exploited. Future research should be able to address the most efficient and validated platform, the most reliable biomarkers that help to select appropriate patients, the most reliable fluid biopsy technique [18], and ways to lower the inherent cost to make MP affordable, particularly for patients in developing and low-income countries [19].