Research ArticlePercutaneous Microwave Coagulation Therapy: A PromisingTherapeutic Method for Breaking the Barrier of theIntertumor Heterogeneity

Haoyue Guo,1,2 Bin Chen,1 Wei Li,1 Hao Wang,1,2 Sha Zhao,1 Peixin Chen ,1,2

Minlin Jiang,1,2 Lishu Zhao,1,2 Kandi Xu,1,2 Hui Sun,1 Yayi He ,1 and Caicun Zhou1

1Department of Medical Oncology, Shanghai Pulmonary Hospital, Tongji University Medical School Cancer Institute,Tongji University School of Medicine, No. 507 Zhengmin Road, Shanghai 200433, China2Tongji University, No. 1239 Siping Road, Shanghai 200433, China

Correspondence should be addressed to Yayi He; 2250601@qq.com

Received 26 July 2021; Revised 19 October 2021; Accepted 27 October 2021; Published 19 November 2021

Academic Editor: Osamah Ibrahim Khalaf

Copyright © 2021 Haoyue Guo et al. -is is an open access article distributed under the Creative Commons Attribution License,which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Intertumor heterogeneity is common in various cancers and has been widely accepted as the primary cause of the diversity andvariation of the effect of the same treatment on patients with the same type of tumor. Percutaneous microwave coagulationtherapy (PMCT) is a minimally invasive and effective approach for destroying tumors by microwave beam under image guidance,which has been applied in lung cancer. However, no previous study has investigated the capability of PMCT solving intertumorheterogeneity. Here, we performed a component analysis of four lung cancer patients’ tumormicroenvironment (TME) via single-cell RNA sequencing (scRNA-seq) and treated all four cases with PMCT. One patient’s TME could be classified into a hot tumor,mainly proinflammatory cytokines, and T cell infiltration. -e other three patients’ TMEs were cold tumors, where immu-nosuppressive cells occupied a large proportion, including tumor-associated macrophages and cancer cells. Despite a high level ofheterogeneity among their tumor microenvironment compositions, disease type and stage, and basic physical conditions, all fourpatients presented a stable disease (SD) without any cancer cell detected in the TME of cancer tissues after PMCT. In conclusion,this report uniquely contributed to the knowledge of the PMCTadaptation to tumor heterogeneity.-erefore, PMCTis promisingto demonstrate a stable and robust antitumor efficacy in unresectable lung cancers with various TMEs.

1. Introduction

Lung cancer remains the most frequent cause of cancer-related mortality worldwide [1]. Histologically, two majorclasses are nonsmall cell lung cancer (NSCLC), accountingfor 85% of lung cancer patients, and small cell lung cancer(SCLC) occupying 15% of lung cancer cases [2]. More than 1million new cases emerge each year, more than 1/3 of whichpresent with distant metastasis and only 15% are at earlystages at the time of diagnosis [3]. -erefore, most lungcancer cases are not appropriate for surgical resection, andvarious therapeutic strategies involving chemotherapy, ra-diology, targeted therapy, and immunotherapy have createda new era of lung cancer treatment [4].

Tumor heterogeneity consists of intertumor and intra-tumor heterogeneity, among which intertumor heterogeneityis generally more visible and has been widely accepted as theprimary cause of the diversity and variation of the effect of thesame treatment on patients with the same type of tumor [5].Moreover, the intertumor heterogeneity has posed significantchallenges on current cancer treatment mentioned above [6],and recent studies on uncovering intertumor heterogeneity oflung cancers have raised enormous hope in therapeutic de-cisions [7,8]. Percutaneous microwave coagulation therapy(PMCT), a novel minimally invasive treatment to destroytumors by microwave beam under image guidance, has beengenerally accepted to overcome these challenges.-e previousstudies considered PMCT could induce cancer cell death [9],

HindawiJournal of Healthcare EngineeringVolume 2021, Article ID 7773163, 9 pageshttps://doi.org/10.1155/2021/7773163

enhance antitumor immunity [10], and inhibit the synthesisof DNA, RNA, and protein in cancer cells [11]. To date, noprevious study has investigated the capability of PMCTsolving intertumor heterogeneity.

In this report, we presented four lung cancer cases beingtreated with PMCT in Shanghai Pulmonary Hospital. Im-portantly, we initially exerted single-cell RNA sequencing(scRNA-seq) techniques on patients’ tumor tissues to ex-plore the composition of their tumor microenvironment(TME). -us, this report uniquely contributed to theknowledge of the PMCT adaptation to intertumor hetero-geneity. -e study was approved by the ethics/licensingcommittee of Shanghai Pulmonary Hospital (approvalnumber: L20-244). We present the following case in ac-cordance with the CARE reporting checklist.

2. Methods

-e fresh tumor tissue was placed in the cold MACS TissueStorage Solution (Miltenyi Biotec, Cat: 130-100-008) andwas transported to the laboratory at low temperature.GEXSCOPETM Tissue Dissociation Solution (Singleron,China) was used to lysate tissues and digest the cells forsingle cell suspension, while the Illumina HiSeq X platform(Illumina, San Diego, CA, USA) was used for sequencing.

3. Case Presentation

Here, we selected four patients who underwent a PMCTbetween March to July, 2019, at the Department of MedicalOncology, Shanghai Pulmonary Hospital. -e follow-updeadline was October, 2019, and the clinical efficacy wasdetermined by checking the CT results from the medicalrecord.

3.1. Case 1. A 66-year-old man (Patient number: H81)presented with a one-month history of left pulmonarynodules and was admitted to our department on July 25,2019. His medical history included the sleeve resection of theleft lower lobe (squamous cell carcinoma, SCC; Figure 1(a),T2N1M0, stage IIB) on September 29, 2016, and fourchemotherapy (gemcitabine) from November 10, 2016, toFebruary 18, 2017 (Figure 2). He was a never-smoker, andhis family history had no history of lung cancer. On pre-sentation, his vital signs were all in normal limitations.Additional remarkable laboratory testing is also shown inTable 1. -e chest computed tomography (CT; June 28,2019) revealed an increased size of left nodules than previousones (Figure 3(a)). Moreover, the pathological and immu-nohistochemical images confirmed relapsed SCC(Figure 1(b)). We performed the scRNA-seq (performed bythe Singleron Biotechnologies) to analyze the composition ofthe patient’s TME, including cancer cells, macrophages,dendritic cells, alveolar cells, and plasma cells (Figure 4(a).-e tumor sample we obtained via CT guided percutaneouslung biopsy (CT-PLB) before the PMCT (July 25, 2019)demonstrated that tumor-associated macrophages (TAMs,CD68+CD163+MRC1+; 33.24%), cancer cells(EPCAM+NAPSA+ SOX2+; 32.20%), and dendritic cells 2

(cDC2, CD1C+FCER1A+CLEC10A+; 22.56%) occupiedthe main component of the TME (Figures 4(b) and 4(c)).

-e patient underwent the PMCT on July 25, 2019. -eappropriate puncture needle was placed into the lesion sitewith the assistance of chest CT, following the skin disin-fection and local anesthesia (2% lidocaine 5mL). -reepieces of tissue were obtained using a biopsy needle. Af-terward, the PMCT puncture needle was replaced into thetumor site, and the heating was controlled as 50w for 3mins(Figure 3(b)). -e immediate chest CT was performed toevaluate the efficacy. Next, we repeated the heating as 50wfor 3mins and performed the chest CT indicating an exu-dation with a diameter of 3 cm at the tumor site. -e needlewas pulled out under the condition of 30w. Another repeatchest CT after the operation demonstrated pneumothoraxand hemorrhage. Within the next month, the patient did nothave fever or hemoptysis, and the efficacy of the PMCTwasevaluated as SD (Figures 3(c) and 3(d)).

3.2. Case 2. A 66-year-old woman (Patient number: he_78)presented with paroxysmal left chest pain for two monthsand visited our hospital on May 14, 2019 (Figure 2). Her pastmedical history was uterine leiomyomatosis and type 2diabetes. She denied any tobacco or alcohol use, and herfamily history was not notable for lung cancers. On physicalexamination, the patient presented with typical vital signs.Further, the laboratory results of he_78 were unremarkable,except for a raised blood glucose level and an amylase level(Table 1). According to pathological examinations, the pa-tient was diagnosed with left pulmonary adenocarcinoma(ADC; TT1N0M0, stage I). -e single-cell sequencingdemonstrated that in the TME before the PMCT (May 14,2019), TAMs accounted for a notable proportion (75.54%),followed by TILs (CD2+CD3D+TRBC2+; 11.96%), cancercells (4.89%), and alveolar type II cells (AT2,EPCAM+SFTPC+ SFTPB+; 4.89%) (Figures 4(b) and 4(c)).

-e patient got PMCT on May 22, 2019. -e process ofthe PMCT was the same as described above (Figure 5(a)),and the H&E staining demonstrated the NSCLC tissue withfew lymphocytes (Figure 1(c)). -e immediate CT after theoperation ruled out the common complications, especiallypneumothorax (Figure 5(b)). Within the next month, thepatient did not have fever or hemoptysis and the efficacy ofthe PMCT was evaluated as SD. -e repeated pathologicalexaminations on June 28, 2019, did not detect any lungcancer tissues (Figure 1(d)).

3.3. Case 3. A 63-year-old man (Patient number: he_57)presented with a 3-year history of relapsed lung cancer andvisited the outpatient on April 18, 2018. He had a back-ground of radical resection of lung cancer under video-assisted thoracoscopic surgery (VATS) on September 11,2015 (adenosquamous carcinoma), right thoracoscopicwedge resection on January 11, 2018 (SCC; Figure 2). He wasan ex-smoker of 40 years with a one-pack day history. Hisfamily history was unremarkable for lung cancers, and vitalsigns on arrival were also unremarkable. Among this pa-tient’s laboratory workup, several indexes showed a

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2015 2016 2017 2018 2019

September 11, 2015

September 29, 2016 November 10, 2016 November 22, 2016

Radical resection of lungcancer under VATS

Sleeve resection ofthe left lower lobe

1st chemotherapy(Gemcitabine)

2nd chemotherapy(Gemcitabine)

3rdchemotherapy(Gemcitabine)

4th chemotherapy(Gemcitabine)

January 6, 2017

January 11, 2018

October 23, 2018 December 25, 2018

June 28, 2019 July 25, 2019

May 22, 2019May 15, 2019

May 9, 2019 May 17, 2019

April 10, 2019

January 9, 2019 February 13, 2019

April 18, 2019 April 19, 2019

February 18, 2017

Right thoracoscopicwedge resection

Chest CT and PET-CT revealed apossibility of left lower lobe ADC

Pathological imagesconfirmed ADC

The pathological imagesconfirmed ADC

The pathological imagesconfirmed SCLC

PMCT

PMCT

PMCT

PMCT

The increased lesionfrom chest CT

The increased lesionfrom chest CT

Pemetrexeddisodium

A possibility of rightresidual lung malignant

Increased size of leftnodules from chest CT

H81

he_78

he_57

he_80

Figure 2: -e timeline of four patients’ lung cancer diagnosis and treatment. VATS, video-assisted thoracoscopic surgery; CT, computedtomography; PET-CT, positron emission tomography-computed tomography; PMCT, percutaneous microwave coagulation therapy; ADC,adenocarcinoma; SCLC, small cell lung cancer.

(a) (b) (c) (d)

(e) (f ) (g) (h)

Figure 1: Hematoxylin and eosin (H&E) staining of the lung biopsy specimen of four cases. -e H＆E staining of H81, whose biopsyspecimen obtained on 29 September 2016 (a) and 29 July 2019 (b), revealed a moderately differentiated squamous cell carcinoma (SCC) withnecrosis, where numerous lymphocytes also infiltrated. -e H＆E staining of he_78, whose biopsy specimen obtained on 23 May 2019 (c),indicated a nonsmall cell lung cancer (NSCLC) with few infiltrating lymphocytes, while that obtained on 28 June 2019 (d) revealed a necrotictissue without any tumor cell or lymphocyte.-e H＆E staining of he_57, whose biopsy specimen obtained on 18 April 2019 (e) and 28May2019 (f), demonstrated a tissue of small cell lung cancer (SCLC) (e) and spindle cell poorly differentiated carcinoma (f), both of which werewithout lymphocyte infiltration. -e H＆E staining of he_80, whose biopsy specimen obtained on 21 May 2019 (g) and 29 July 2019 (h),displayed a micropapillary lung adenocarcinoma (g) and ductal type adenocarcinoma (h), both of which were with little lymphocyteinfiltration.

Table 1: Basic clinical information and abnormal laboratory results of the four patients in this report.

Patientnumber Age/gender Stage Pathology Location of

lesionsAbnormal laboratory results

Index Value Reference range

H81 66 y/male II SCC Left upper lobe

Monocyte percentage 0.09% 3.0%–8.0%Eosinophilic granulocyte percentage 0.05% 1.0%–5.0%

Basophilic granulocyte count 0.42×109 0.05–0.30×109

INR 1.19 R 0.80–1.10 RTBA 18.5 μmol/L 0–10 μmol/L

Homocysteine 18.1 μmol/L 3–15 μmol/LBlood glucose 3.9mmol/L 4.1–5.9mmol/L

he_78 66 y/female I ADC Left upper lobe Blood glucose 8.1mmol/L 4.1–5.9mmol/LAmylase 119U/L <100U/L

he_57 63 y/male IV SCLC Right lower lobe

Eosinophilic granulocyte percentage 0.80% 1.0%–5.0%Potassium 3.44mmol/L 3.5–5.1mmol/L

Calcium 2.16mmol/L 2.18–2.60mmol/L

Myoglobin 21.00 ng/ml 28–72 ng/ml

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moderate decrease (Table 1). -e positron emission to-mography-computed tomography (PET-CT) revealed apossibility of right residual lung malignant tumors (April 10,2019). On April 18, 2019, he revisited Shanghai PulmonaryHospital, and the pathological and immunohistochemicalresults (Figure 1(e)) via biopsy confirmed the right small celllung cancer (SCLC; T1N0M1, stage IV). As for the com-position of the TME, TILs (78.57%), TAMs (10.12%), AT2(8.93%), and cDC2 (2.38%) were all detected components(Figures 4(b) and 4(c)).

After a discussion with the radiology team of treatmentoptions, we decided to exert the PMCT on April 19, 2019.-e procedure of the PMCT was the same as describedabove (Figure 6(a)). -e immediate CT after the operationruled out the common complications, especially pneu-mothorax (Figure 6(b)). Within the next month, the patientdid not have fever or hemoptysis, and the efficacy of thePMCT was evaluated as SD (Figure 6(c)). During the

period, another repeated pathological examination de-tected a tissue of spindle cell poorly differentiated carci-noma (Figure 1(f )).

3.4. Case 4. An 80-year-old man (Patient number: he_80)presented with the CT finding of lung cancer for fourmonths and came to our hospital on May 16, 2019. -epatient initially took a medical examination due to lumbagoon October 23, 2018, and the chest CTand PET-CTrevealeda possibility of left lower lobe ADC.-e pathological imageson December 25, 2018, confirmed ADC (T3N2M0, stageIIIc). -erefore, he was started on the pemetrexed disodiumon January 9, 2019, without distinct adverse reactions(Figure 2). Chest CT (February 13, 2019, and May 9, 2019)showed the increased lesion sites, and the moderate en-largement of subpleural lesions in the left lower lobe(Figure 7(a)). Additionally, the patient had a background of

Table 1: Continued.

Patientnumber Age/gender Stage Pathology Location of

lesionsAbnormal laboratory results

Index Value Reference range

he_80 80 y/male III ADC Left lower lobe

Hemoglobin 99.0 g/l 130–175 g/lRed cell count 3.48×1012/L 4.3–5.8×1012/L

Neutrophil percentage 0.43% 50.0–70.0%Lymphocyte percentage 0.44% 20.0–40.0%Monocyte percentage 0.10% 3.0–8.0%

D-Dimer 268 ng/ml 0–232 ng/mlBlood glucose 7.0mmol/L 4.1–5.9mmol/L

Oxygen partial pressure 70.3mmHg 80–100mmHgTotal hemoglobin 10.8 g/dL 12–17.5 g/dLOxygen saturation 0.948 96–100%Oxyhemoglobin 0.925 94–99%Methemoglobin 0.009 0.2–0.6%

Standard alkali remaining 3.4mmol/L −3–3mmol/LAlkali remaining 3.3mmol/L −3–3mmol/L

Bicarbonate 27.4mmol/L 22–27mmol/LStandard bicarbonate 27.3mmol/L 22–27mmol/L

Alveolar-arterial partial pressuredifference 31.5mmHg 8–25mmHg

SCC, small cell lung cancer; ADC, adenocarcinoma; SCLC, small cell lung cancer; INR, international normalized ratio; TBA, total bile acids.

(a) (b) (c) (d)

Figure 3: CTimages of He_81. (a)-emass in the left upper lobe (June 28, 2019). (b)-e process of PMCT (July 25, 2019). (c) Follow-up CTshowing a SD to PMCT on August 27, 2019. (d) Follow-up CT showing a SD to PMCT on October 29, 2019. CT, computed tomography;PMCT, percutaneous microwave coagulation therapy; SD, stable disease.

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emphysema, bullae, lacunar infarction, and benign prostatichyperplasia. He denied any tobacco or alcohol use, withoutany notable family history of lung cancer. -e patient’s bloodpressure, heart rate, respiratory rate, and temperature were allin normal limitations. However, due to his advanced age, thispatient had a series of abnormalities in the laboratory ex-amination (Table 1). Importantly, the pulmonary functiontest of this patient showed a moderately decreased lungventilation, and the electrocardiogram result demonstratedatrial premature beats and the complete right bundle branchblock. -e single-cell sequencing proved that cancer cellsaccounted for the most significant proportion of the TME

(84.54%), followed by AT2 (6.91%), TAMs (4.28%), TILs(3.29%), cDC2 (2.72%), and plasma (0.33%; Figures 4(b) and4(c)).

-e patient underwent the PMCTon May 17, 2019, withthe same procedure as mentioned above (Figure 7(b)), andthe pathological biopsy indicated ADC (Figure 1(g)). -eimmediate CT after the operation ruled out the commoncomplications, especially pneumothorax. Within the nextmonth, the patient did not have fever or hemoptysis, and theefficacy of the PMCT was evaluated as SD (Figure 7(c)).During the period, a repeated pathological examinationconfirmed ADC (Figure 1(h)).

Cancer cell (EPCAM+NAPSA+SOX2+)Macrophage (CD68+CD163+MRC1+)cDC2 (CD1C+FCER1A+CLEC10A+)T cell (CD2+CD3D+TRBC2+)Plasma cell (CD79A+IGHG1+JCHAIN+)AT2 (EPCAM+SFTPC+SFTPB+)

-25 0 25tSNE_1

tSN

E_2

20

0

-20

-40

(a)

-25 0 25tSNE_1

tSN

E_2

20

0

-20

-40

H81 (total cell number: 3823)he_57 (total cell number: 286)he_78 (total cell number: 713)he_80 (total cell number: 586)

(b)

he_80

he_78

he_57

H81

Sam

ple

0.00 0.25 0.50 0.75 1.00Percent

AT2Cancer cellcDC2

MacrophagePlasmaT cell

(c)

Figure 4:-e quantitative and functional analysis of tumormicroenvironment (TME) via single-cell RNA sequencing (scRNA-seq). (a)-etSNE plot shows the cell types we detected in this study. Different colors represent different cell types. (b)-e tSNE plot shows the source ofsamples. Different colors represent different patients. (c) -e proportion of different types of cells in each sample. In the sample of he_80,84.54% are cancer cells, 4.28% are macrophages, 0.66% are cDC2s, 3.29% are T cells, 0.33% are plasma cells, and 6.91% are AT2s. In thesample of he_78, 10.12% are macrophages, 2.38% are cDC2s, 78.57% are T cells, and 8.93% are AT2s. In the sample of he_57, 4.89% arecancer cells, 75.54% are macrophages, 2.72% are cDC2s, 11.96% are T cells, and 4.89% are AT2s. In the sample of H81, 32.20% are cancercells, 33.24% are macrophages, 22.56% are cDC2s, 7.28% are T cells, 4.18% are plasma cells, and 0.54% are AT2s. tSNE, t-distributedstochastic neighbor embedding; cDC2s, dendritic cells 2; AT2s, alveolar type II cells.

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Cold tumorHot tumor

Extra-cellularmatrix

Cancer-associatedfibroblast

Pericyte

Red blood cell

Patient number:he_57

Lengends

Macrophage Dendritic cell Myeloid-derivedsuppressor cell

T lymphocyte

Alveolar typeII cells

Cancer cellNatural killer

cell

Patient number:he_80

Patient number:he_78

Patient number:H81

Figure 5: CT images of he_78. (a) -e process of PMCT (May 22, 2019). (b) -e immediate CT after the operation (May 23, 2019). (c)Follow-up CT showing a SD to PMCT (July 1, 2019). CT, computed tomography; PMCT, percutaneous microwave coagulation therapy.

(a) (b) (c)

Figure 6: CT images of he_57. (a) -e process of PMCT (April 19, 2019). (b) -e CTafter the operation (April 29, 2019). (c) Follow-up CTshowing a SD to PMCT on May 21, 2019. CT, computed tomography; PMCT, percutaneous microwave coagulation therapy.

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4. Discussion

PMCT is a noninvasive and effective approach for lungcancers [12,13], which has been proven to produce tissueheating effects in a much broader range than radiofrequencyablation [14]. However, few data revealed the effectiveness ofPMCT in treating lung cancer with different types of TME.In this report, we presented four cases with various lungcancers, ADC, SCC, and SCLC, and a wide range of agesfrom 63 to 80 years, which were treated with PMCT andpresented SD. Moreover, the post-PMCTscRNA-seq resultsof these four patients were all no cancer cells, demonstratingthat most of the cancer cells in the residual tumor lesionsmight be necrotic. Among these patients, only one patientpresented pneumothorax, while the rest did not develop anycomplications.

As is well known, the TME is essential for tumorigenesisand disease progression, which consists of various innateand adaptive immune cells, stromal cells, blood vessels, andsecreted factors [15]. However, the high level of heteroge-neity within the TME hinders the development of lungcancer research and treatment. -erefore, finding out the

composition, quantity, and function of cancer-infiltratingimmune cells is the crux for optimizing therapeutic plans.-e TME is generally classified into two types: “cold tumor”with mainly immunosuppressive cells (TAMs, myeloid-derived suppressor cells, regulatory T cells, etc.) and “hottumor” with mainly proinflammatory cytokines and T cellinfiltration (Figure 8) [16]. What is more, a hot tumor isconsidered a better response to immunotherapy [17]. In thisreport, the TME of four patients could also be divided intocold tumors (H81, he_78, and he_80) and hot tumor(he_57), and all cases responded well to PMCTregardless ofthe TME class. -ence, we proved the vast potential ofPMCT to address the cold tumor-associated challenges ofimmunotherapy.

Secondly, aging is another major challenge for cancertreatment. Approximately 50% of lung cancer cases are 70years old or older, and about 14% are over 80 years old [18].However, according to a national analysis in the US, 57.6%of patients over 90 years old did not receive any treatmentbetween 2004 and 2014, out of concerns about their toler-ance and quality of life [19]. In this case, local treatment hasshown a steep increase in lung cancer patients over 80 [20].

(a) (b) (c)

Figure 7: CT images of he_80. (a)-e mass in the left lower lobe (May 9, 2019). (b)-e process of PMCT (May 17, 2019). (c) Follow-up CTshowing a SD to PMCT on July 4, 2019. CT, computed tomography; PMCT, percutaneous microwave coagulation therapy; SD, stabledisease.

(a) (b) (c)

Figure 8: -e major classifications of tumor microenvironment (TME). Generally, TME can be divided into cold tumors and hot tumors.Hot tumors are characterized as sufficient tumor-infiltrating T lymphocytes while cold tumors are characterized as adequate immuno-suppressive cells, including tumor-associated macrophages, myeloid-derived suppressor cells, and regulatory Tcells. In this report, the TMEof he_57 can be classified into a hot tumor, and H81, he_78, and he_80 can be classified into cold tumors based on the componentsmentioned in Figure 2.

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In this report, patient 4 (he_80) is an 80-year-old patientwith poor cardiopulmonary function and basic physicalcondition. Surprisingly, this patient did not show prominentadverse events and discomfort after PMCTwith an excellentcurative effect, which offers a safe and justifiable perspectivefor future elderly lung cancer patients.

Besides, the tumor lesion after PMCT is kept in the lungtissue rather than being resected. -erefore, the tumor sizeof four patients we reported was not a sensitive criterion forcurative effect evaluation of PMCT.

-ere remain several drawbacks of this report thatshould be mentioned. First, all of the samples we obtained toperformwere from biopsy, with high preference and locality.-e total cell numbers of samples from he_57, he_78, andhe_80 are relatively low, so it may not fully reflect all thecomponents of their TMEs. Secondly, we tried to obtain lungcancer tissues of four patients repeatedly after the PMCT butfailed, since all of the samples were not detected with anycancer cell. In future studies, we will continue to explore theeffect of PMCTon the TME by comparing the TME of pre-and post-PMCT patients. -ird, the follow-up time for theabove patients is one month, and we will continue to tracktheir disease and survival in the future. Last, no tumor cellswere found in the pathological examination after the PMCT,so no further scRNA-seq were carried out.

In conclusion, PMCT is a safe and effective method forlung cancers with various TMEs, and more prospectiveresearch on a larger scale with long-term follow-up isneeded.

Data Availability

All data are presented in the article.

Ethical Approval

-e authors are accountable for all aspects of the work inensuring that questions related to the accuracy or integrity ofany part of the work are appropriately investigated andresolved. All procedures performed in the studies involvinghuman participants were in accordance with the ethicalstandards of the institutional and/or national researchcommittee(s) and with the Helsinki Declaration (as revisedin 2013). Written informed consent was obtained from thepatients.

Consent

Written informed consent was obtained from all thepatients.

Conflicts of Interest

-e authors declare that they have no conflicts of interest.

Authors’ Contributions

Haoyue Guo, Bin Chen, and Wei Li contributed equally tothis work.

Acknowledgments

-is study was supported in part by a grant of NationalNatural Science Foundation of China (81802255), ClinicalResearch Project of Shanghai Pulmonary Hospital(FKLY20010), Young Talents in Shanghai (2019 QNBJ),‘Dream Tutor’ Outstanding Young Talents Program(fkyq1901), Clinical Research Project of Shanghai Pulmo-nary Hospital (FKLY20001), Respiratory Medicine, a KeyClinical Specialty Construction Project in Shanghai, Pro-motion, and Application of Multidisciplinary CollaborationSystem for Pulmonary Noninfectious Diseases, ClinicalResearch Project of Shanghai Pulmonary Hospital(fk18005), Key Discipline in 2019 (oncology), Project ofShanghai Municipal Health Commission (201940192),Project of Shanghai Municipal Science and TechnologyCommission , Scientific Research Project of Shanghai Pul-monary Hospital (fkcx1903), Shanghai Municipal Com-mission of Health and Family Planning (2017YQ050),Innovation Training Project of SITP of Tongji Universityand Key Projects of Leading Talent (19411950300), andYouth Project of Hospital Management Research Fund ofShanghai Hospital Association (Q1902037). -e authorswant to thank Singleron for providing single-cell detectionand data analysis and thank BioRender (https://app.biorender.com/) for providing them a platform to createFigure 8 with a legal authorization for publication.

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